Angiopoietin-2 predicts mortality in male ESKD patients on hemodialysis
Individuals suffering from end-stage kidney disease (ESKD) undergoing dialysis have significantly higher mortality rates compared to patients with non-ESKD of the same age. Approaches to assess mortality risk, which could enable tailored and effective care are needed (1).
Angiopoietin-2 (Ang2) is a protein that is crucial in pathological vascular restructuring and in the formation of new blood vessels (angiogenesis). These processes are active in ESKD patients and might contribute to the elevated mortality observed in this patient group.
Angiopoietin-2 predicts mortality in male ESKD patients on hemodialysis
In a recent multicenter prospective study Ang2 was measured in 340 hemodialysis (HD) patients with ESKD to predict overall mortality (2, 3). The patients were followed up for 5-years and blood samples and clinical data were collected at baseline. Ang2 serum levels were measured with a enzyme-linked immunosorbent assay (ELISA) from Biomedica.
Angiopoietin-2 predicts mortality in male ESKD patients on hemodialysis
The results of the study demonstrated that the risk of death was higher in male patients with elevated Ang2 levels than in female patients: Ang2 levels at baseline are independently associated with all cause mortality in male ESKR patients on HD.
Angiopoetin-2 can reliably be measured in serum or plasma samples with a conventional ELISA assay:
Chronic Kidney Disease (CKD) affects more than 10% of the general population worldwide (1). It is characterized by kidney fibrosis, a progressive process that destroys the kidney (2). An effective treatment to stop the progression of CKD is still lacking. Scientists have now discovered a potential mean to treat CKD by inhibiting Angiopoietin-2 (ANG2), a vascular growth factor (3). Angiopoietin-2 is a potential new treatment target for kidney fibrosis in CKD.
Angiopoietin-2 a new treatment target for kidney fibrosis in CKD
Patients with chronic kidney disease (CKD) have increased plasma levels of Angiopoietin-2 (ANG2). Studies have shown that ANG2 induces kidney injury and fibrosis in patients with CKD as well as in mouse models of kidney disease (3). Preventing the progression of renal fibrosis by blocking Angiopoietin-2 could be a new approach in treating chronic kidney disease-associated pathologies.
Based on data of the recent study from Chang et al., on the application of an Angiopoietin-2 inhibitor to mouse models of progressive kidney disease, the scientists found a profound inhibitory effects of ANG2 inhibition on macrophage infiltration and fibrosis (3).
Plasma levels of angiopoietin-2 are increased in patients with chronic kidney disease (CKD). Moreover, mouse models of progressive kidney disease also demonstrate increased angiopoietin-2 in both plasmas and kidneys. The role of dysregulated angiopoietins in the progression of kidney disease has not been thoroughly investigated. Here, we found in a cohort of 319 patients with CKD that plasma angiopoietin-2 and angiopoietin-2/angiopoietin-1 ratios were positively associated with the development of kidney failure. In mice with progressive kidney disease induced by either ureteral obstruction or ischemia-reperfusion injury, overexpression of human angiopoietin-1 in the kidney tubules not only reduced macrophage infiltration in the initial stage post-injury but also attenuated endothelial cell apoptosis, microvascular rarefaction, and fibrosis in the advanced disease stage. Notably, angiopoietin-1 attenuated chemokine C-C motif ligand 2 (CCL2) expression in the endothelial cells of the fibrosing kidneys, and these protective effects led to attenuation of functional impairment. Mechanistically, angiopoietin-1 reduced CCL2-activated macrophage migration and protected endothelial cells against cell apoptosis induced by angiopoietin-2 and Wnt ligands. Based on this, we applied L1-10, an angiopoietin-2 inhibitor, to the mouse models of progressive kidney disease and found inhibitory effects on macrophage infiltration, microvascular rarefaction, and fibrosis. Thus, we defined the detrimental impact of increased angiopoietin-2 on kidney survival of patients with CKD which appears highlighted by angiopoietin-2 induced endothelial CCL2-activated macrophage infiltration and endothelial cell apoptosis in their kidneys undergoing fibrosis.
Baseline Ang-2 plasma levels are an independent prognostic biomarker in refractory metastatic colorectal cancer
Chemorefractory is a term that is used to describe a cancer that does not respond to chemotherapy. Currently, no biomarkers are available to predict the efficacy of chemotherapy in chemorefractory metastatic colorectal cancer. Researchers from Italy have shown that circulating biomarker levels of Angiopoietin-2 (ANG2) increases early and predicts outcome with regorafenib but not with trifluridine/tipiracil treatment. Thus, baseline Ang-2 plasma levels are an independent prognostic biomarker in chemorefractory metastatic colorectal cancer. Read more: Early modulation of Angiopoietin-2 plasma levels predicts benefit from regorafenib in patients with metastatic colorectal cancer
Easy measurement of Angiopoietin-2 (ANG-2) in blood samples with only 20µl sample volume.
Antoniotti C, Marmorino F, Boccaccino A, Martini S, Antista M, Rossini D, Zuco V, Prisciandaro M, Conca V, Zucchelli G, Borelli B, Cosentino P, Germani MM, Bosco MF, Carullo M, Vetere G, Moretto R, Giordano M, Masi G, Pietrantonio F, Zaffaroni N, Cremolini C. Eur J Cancer. 2022. 165:116-124. doi: 10.1016/j.ejca.2022.01.025. PMID: 35231767.
Abstract
Background: No biomarkers are currently available to predict the efficacy of trifluridine/tipiracil (FTD/TPI) in chemorefractory metastatic colorectal cancer. The multicohort REGOLAND study aims at exploring and validating circulating markers potentially able to predict benefit from regorafenib in this setting. Material and methods: In the retrospective ‘regorafenib exploratory cohort’, including 105 patients treated with regorafenib, baseline (d1) plasma levels of angiogenesis-related biomarkers and their early modulation after 15 days (d15) of treatment were investigated for correlation with clinical outcome. Based on a pre-specified statistical hypothesis, main retrospective findings were prospectively challenged in the ‘regorafenib validation cohort’, including 100 patients treated with regorafenib. Prospectively validated putative biomarkers were then assessed in the control ‘FTD/TPI cohort’, including 93 patients treated with FTD/TPI. Results: In the ‘regorafenib exploratory cohort’, the early (d15) increase of Angiopoietin-2 (Ang-2) was associated with longer progression-free survival (HR:0.57 [95%CI:0.38-0.88], P = 0.004) and a trend towards longer OS (HR:0.74 [95%CI:0.48-1.14], P = 0.165), than the early decrease. Similar results were prospectively confirmed in the ‘regorafenib validation cohort’ (HR for progression-free survival:0.72 [95%CI:0.48-1.08], P = 0.095; HR for OS:0.77 [95%CI:0.51-1.16], P = 0.204). No predictive impact was shown for the early modulation of Ang-2 in the ‘FTD/TPI cohort’. High baseline Ang-2 levels predict poor prognosis in all the investigated cohorts, independently of other clinical prognostic variables. Conclusions: The early modulation of circulating Ang-2 predicts the efficacy of regorafenib. Baseline Ang-2 plasma levels are an independent prognostic biomarker in chemorefractory metastatic colorectal cancer.
Munakata S, Ueyama T, Ishihara H, Komiyama H, Tsukamoto R, Kawai M, Takahashi M, Kojima Y, Tomiki Y, Sakamoto K. J Gastrointest Cancer. 2021.52(1):237-242. PMID: 32166589.
Circulating Angiopoietin-2 (ANG2) increases with PAP sleep apnea treatment:Obstructive sleep apnea (OSA), the most common sleep-related breathing disorder, is becoming increasingly prevalent worldwide due to widespread obesity. It causes a person to repeatedly stop and start breathing during sleep. Positive airway pressure (PAP) is a well-established treatment for OSA patients, using a machine to pump air under pressure into the airway of the lungs. Though effective, the therapy causes large increases in lung volumes during the night that are potentially deleterious. A recent study investigated the impact of PAP therapy on various biomarkers related to alveolar epithelial and endothelial injury. The researchers have shown that Angiopoietin-2 (ANG-2), a marker of endothelial injury and cardiovascular disease risk, increases with continuous positive airway pressure therapy . This finding raises concern for a possible adverse impact of PAP therapy on the vascular endothelium.
Gottlieb DJ, Lederer DJ, Kim JS, Tracy RP, Gao S, Redline S, Jelic S. Sleep Med. 2022 May 16;96:119-121. doi: 10.1016/j.sleep.2022.05.007. Epub ahead of print. PMID: 35636149.
Abstract
Background: Obstructive sleep apnea (OSA) has been identified as a possible contributor to interstitial lung disease. While positive airway pressure (PAP) is effective therapy for OSA, it causes large increases in lung volumes during the night that are potentially deleterious, analogous to ventilator-induced lung injury, although this has not been previously studied. The goal of this study was to assess the impact of PAP therapy on four biomarkers of alveolar epithelial and endothelial injury and extracellular matrix remodeling in patients with OSA.
Methods: In 82 patients with moderate to severe OSA who were adherent to PAP therapy, surfactant protein D, osteopontin, angiopoietin-2, and matrix metalloprotease-7 were measured by ELISA in serum samples collected before and 3- to 6-months after initiation of PAP therapy.
Results: An increase in angiopoietin-2 level of 0.28 ng/mL following PAP therapy was observed (p = 0.007). This finding was replicated in an independent sample of OSA patients. No significant change was detected in surfactant protein D, osteopontin, or matrix metalloprotease-7.
Conclusions: This finding raises concern for a possible adverse impact of PAP therapy on vascular endothelium.
Angiopoietin-2 can easily be measured with an ELISA assay in serum and plasma samples with only 20 µl of sample volume! The kit incorporates ready to use standards and controls. The assay range is optimized for clinical samples- no sample dilution required!
√ Immediate results: no sample dilution required √ Full validation package – the assay is optimized for clinical samples √ Kit includes ready to use standards and controls √ HIGH QUALITY GUARANTEED
Al-Terki A, Abu-Farha M, AlKhairi I, Cherian PT, Sriraman D, Shyamsundar A, Ali S, Almulla F, Tuomilehto J, Abubaker JA. Front Endocrinol (Lausanne). 2018 Nov 13;9:651. doi: 10.3389/fendo.2018.00651. PMID: 30524367; PMCID: PMC6262344.
Abstract
Objective: Obstructive sleep apnea (OSA) is a sleep disorder caused by the complete or partial obstruction of the upper airways. The worldwide prevalence of OSA is increasing due to its close association with obesity epidemic and multiple health complications, such as hypertension, cardiovascular disease, and Type 2 diabetes. Angiopoietin-like protein (ANGPTL)-4 and ANGPTL8 (betatrophin) have been suggested to play a role in the development of these diseases through their role in regulating the metabolism of plasma lipid molecules. This study was designed to evaluate ANGPTL4 and 8 levels in an OSA group and a control group to clarify the effect of OSA on ANGPTL4 and 8 levels. Methods: In total, 74 subjects were enrolled in this study, including 22 age- and body mass index (BMI)-matched controls with the Apnea Hypopnea Index (AHI) score of <5 events/h and 52 subjects with an AHI score of >5 events/h. Sleep apnea was assessed using a portable sleep test. ANGPTL4 and 8 levels were measured in plasma samples using enzyme-linked immunosorbent assay. Results: Mean AHI score (2.5 ± 1.6) in the control group was significantly lower than that in the OSA group (22.9 ± 17.9; p < 0.0001). Leptin, interleukin-(IL) 6, insulin, and HOMA-IR values were higher in the OSA group than in the control group. ANGPTL8 level was higher in the OSA group (1130.0 ± 108.61 pg/mL) than in the control group (809.39 ± 108.78 pg/mL; p = 0.041). Similarly, ANGPTL4 was higher in the OSA group (179.26 ± 12.89 ng/mL) than in the control group (142.63 ±7.99 ng/mL; p = 0.018). Conclusion: Our findings demonstrate that ANGPTL4 and 8 levels were increased in subjects with OSA, suggesting that the upregulation of these lipid metabolism regulators might play a role in lipid dysregulation observed in people with OSA.
Gao S, Emin M, Thoma T, Pastellas K, Castagna F, Shah R, Jimenez A, Patel N, Wei Y, Jelic S. Sleep. 2021 Apr 9;44(4):zsaa286. doi: 10.1093/sleep/zsaa286. PMID: 33351148; PMCID: PMC8033461.
Abstract
Study objective: Obstructive sleep apnea (OSA) is highly prevalent and triples vascular thromboembolic risk. Intermittent hypoxia (IH) during transient cessation of breathing in OSA impairs endothelial protection against complement. Complement activation stimulates the endothelial release of a pro-thrombotic von Willebrand factor (vWF). We investigated whether increased complement activity in OSA promotes the endothelial release of vWF and pro-inflammatory angiopoietin-2. We further investigated whether improving complement protection with statins reverses these changes.
Methods: Using endothelial cells (ECs) and blood collected from OSA patients (n = 109) and controls (n = 67), we assessed whether altered cellular localization of complement inhibitor CD59 in OSA modulates exocytosis of Weibel-Palade bodies (WPB), secretory granules that store vWF and angiopoietin-2. These interactions were also assessed in vitro in ECs exposed to normoxia or IH with or without recombinant complement C9 and with or without atorvastatin.
Results: Circulating levels of angiopoietin-2 were greater in OSA than controls and levels of vWF cleavage products correlated with OSA severity. In cultured ECs, IH enhanced complement-stimulated angiopoietin-2 and vWF release by reducing EC surface and increasing intracellular expression of complement inhibitor CD59. Intracellular CD59 co-localized with WPB in OSA. IH increased binding of intracellular CD59 to syntaxin-3, which dissociated syntaxin-3 from voltage-sensitive calcium channel Cav1.2, and activated WPB exocytosis in a calcium-dependent manner. Atorvastatin reversed IH-enhanced endothelial release of vWF and angiopoietin-2.
Conclusions: IH promotes the complement-mediated release of vWF and angiopoietin-2, which may contribute to pro-thrombotic and pro-inflammatory conditions in OSA. Statin reversed these effects, suggesting a potential approach to reduce cardiovascular risk in OSA.
#Acute pulmonary embolism (PE) is a common and potentially fatal disease. The cause is usually a blood clot in the leg that travels to the lung and blocks blood flow . #Angiopoietin-2 (Ang-2) is a signaling molecule that is upregulated in response to insufficient tissue oxygen supply. In a recent study scientists investigated the potential role of Ang2 as a #risk factor in patients with acute PE. The results indicate that Ang2 correlates with PE severity, right ventricular dysfunction, and need for #ICU admission. Thus, Ang-2 holds promise as a novel marker that can aid in risk stratification for this patient population. Learn more https://lnkd.in/eFvntsA
Biomedica offers an Angiopoietin-2 ELISA Assay Kit that is optimized for clinical samples.
Highlights √ Immediate results – no sample dilution required √ Full validation package – developed and manufactured by Biomedica √ Kit includes ready to use standards and controls √ Automatable √ HIGH QUALITY GUARANTEED – https://buff.ly/3etWGHH
The proinflammatory cytokine Angiopoietin-2 plays a key role in endothelial cell disruption and associated events. A recent study demonstrates that a three day change in Angiopoeitin-2 levels predicts the clinical course in hospital-mortality of patients with SARS-CoV-2. Angiopoietin-2 is a relevant part of disease pathogenesis and could be a target for new specific treatments in these patients.
Read more: Dynamic angiopoietin-2 assessment predicts survival and chronic course in hospitalized patients with … Villa E et al., Blood Adv. 2021 Feb 9;5(3):662-673.
Angiopoietin-2 (ANG-2) can reliably be measured by ELISA in human serum, plasma and urine samples with Biomedica’s fully validated assay kit. Only 20 µl of sample volume is required! The kit incorporates ready to use standards and controls. The assay range is optimized for clinical samples- no sample dilution required!
Features & Benefits
√ Immediate results: no sample dilution required √ Full validation package – the assay is optimized for clinical samples √ Kit includes ready to use standards and controls √ Automatable √ HIGH QUALITY GUARANTEED – https://buff.ly/3etWGHH
Studies show that Angiopoietin-2 is a strong predictor in patients with SARS-CoV-2
Two research teams find that increased levels of the proinflammatory cytokine Angiopoietin-2 is crucial to predict transfer to the intensive care units and is responsible for hypercoagulation observed in critically ill patients infected with SARS-CoV-2.
Studies:
Angiopoietin-2 as a marker of endothelial activation is a good predictor factor for intensive care unit admission of … patients. Smadja DM et al., Angiogenesis, 2020;1-10. Full text
Elevated Angiopoietin-2 inhibits thrombomodulin-mediated anticoagulation in critically ill … patients.Hultstrom M et al., MedRxiv preprint server, 2021.Full text
SARS-CoV-2 infection has a diverse range of symptoms and may cause severe illness, in particular in patients with cardiovascular risk factors (1).
The infection is associated with an increase in procoagulant factors that have been shown to be associated with a higher mortality in patients with SARS-CoV-2 pneumonia (2). Pulmonary embolism and microvascular thrombosis in the lungs have been described in several reports in acute respiratory distress syndrome as well as in patients infected with SARS-CoV-2 (3).
Endothelial damage and inflammation in SARS-CoV-2 infection
The inflammatory cytokine storm occurring in patients infected with SARS-CoV-2, leads to the recruitment of leukocytes which damage the capillary endothelium. The disruption of the thromboprotective effect of endothelial cells likely leads to microvascular thrombosis. Thus, the endothelial damage and inflammation in several organs in patients with SARS-CoV-2 infection is a direct consequence of viral involvement and of the host inflammatory response (4).
Despite the routine thrombosis prophylaxis as standard of care treatment, the major complication in patients infected with SARS-CoV-2 is the hyperactivation of the coagulation system indicating a poor prognosis among these patients in intensive care (5).
Angiopoietin-2 (ANG2) is a soluble marker of endothelial activation
Angiopoietin-2 is a soluble protein that is involved in the regulation of vascular integrity, quiescence, and inflammation (6). ANG2 contributes to the formation of new vasculature. During angiogenesis, ANG2 exerts its effects via the angiopoietin-1/TIE2 receptor signaling system on endothelial cells. Disruption of this signaling leads to the loss of endothelial integrity. In consequence, the endothelium responds to various pro-inflammatory cytokines and growth factors.
Thus, Angiopoietin-2 is a soluble marker of endothelial activation and is involved in hemostasis, thrombo-inflammatory events or sepsis (7, 8).
ANG2 is a crucial factor to predict transfer to the intensive care unit
ANG2 induces inflammation and vascular hyperpermeability and correlates with adverse outcomes in several critical care syndromes (9). In patients infected with SARS-CoV-2, ANG2 was recently reported by Smadja and colleagues to be a relevant factor to predict transfer to the intensive care unit as it was associated with poor lung compliance (7). Thus, showing that endothelial activation reinforces the hypothesis of an infection-associated microvascular dysfunction.
Angiopoietin-2 inhibits anticoagulation in critically ill patients infected with SARS-CoV-2
Based on the above findings, Hulstrom and colleagues investigated if the highly elevated inflammatory cytokine ANG2 levels observed in SARS-CoV-2 patients had a direct effect on the coagulation system (10). Animal models and in vitro assays were included in their investigation. As thrombomodulin is expressed on the surface of endothelial cells where it binds thrombin and inhibits the intrinsic coagulation process, the scientists observed that ANG2 inhibits thrombomodulin-mediated anticoagulation and activation of protein C in human donor plasma. Furthermore, the increased ANG2 levels in patients infected with SARS-CoV-2 correlated with disease severity, hypercoagulation, and mortality by promoting hypercoagulation.
Revealing this novel mechanism for ANGPT2 in hypercoagulation the scientists suggest that inhibition of Angiopoietin-2 may be tested for treating severe cases of SARS-CoV-2 infection, as well as in certain other conditions, including sepsis (10).
Angiopoietin-2 (ANG-2) can reliably be measured by ELISA in human serum, plasma and urine samples with Biomedica’s fully validated assay kit. Only 20 µl of sample volume is required! The kit incorporates characterized epitope mapped antibodies and ready to use standards and controls. The assay range is optimized for clinical samples- no sample dilution required!
Features & Benefits
√ CONVENIENT – Assay range optimized for clinical samples;
– Ready to use standards and 2 controls included
√ RELIABLE – Rigorously validated according to FDA/EMEA/ICH guidelines
√ SPECIFIC – Characterized, epitope mapped capture and detection antibodies
√ QUALIFIED CUSTOMER SERVICE – supports you every step of the way
Clinical course and risk factors for mortality of adult inpatients with … in Wuhan, China: a retrospective cohort study. Zhou F et al., Lancet, 2020; 395(10229):1054-1062. https://pubmed.ncbi.nlm.nih.gov/32171076/ PMID: 32171076
Abnormal Coagulation parameters are associated with poor prognosis in patients with novel coronavirus pneumonia. Tang N et al., J Thromb Haemost, 2020; 18(4): 844-847. https://pubmed.ncbi.nlm.nih.gov/32073213/ PMID: 32073213
Pulmonary embolism in … patients: awareness of an increased prevalence. Poissy J et al, Circulation, 2020; 14; 142(2):184-186. https://pubmed.ncbi.nlm.nih.gov/32330083/ PMID: 32330083
… and coagulation: bleeding and thrombotic manifestations of SARS-CoV-2 infection. Al-Samkari H et al., Blood, 2020:136, 489-500. https://pubmed.ncbi.nlm.nih.gov/32492712/ PMID: 32492712
Role of Angiopoietin-2 in Vascular Physiology and Pathophysiology. Akwii RG et al., Cells, 2019; 8(5): 471. https://pubmed.ncbi.nlm.nih.gov/31108880/ PMID: 31108880
Angiopoietin-2 as a marker of endothelial activation is a good predictor factor for intensive care unit admission of … patients. Smadja DM et al., Angiogenesis, 2020;1-10. https://pubmed.ncbi.nlm.nih.gov/32458111/ PMID: 32458111
Angiopoietin 2 levels in the risk stratification and mortality outcome prediction of sepsis-associated coagulopathy. Statz S et al., Clin Appl Thromb Hemost 24(8):1223–1233. https://pubmed.ncbi.nlm.nih.gov/29996658/ PMID: 29996658
Circulating angiopoietin-2 and the risk of mortality in patients with acute respiratory distress syndrome: a systematic review and meta-analysis of 10 prospective cohort studies. Li F et al., Therapeutic advances in respiratory disease, 2020; 14, 1753466620905274 (2020). https://pubmed.ncbi.nlm.nih.gov/32043429/ PMID: 32043429
Rigorously validated:the human Angiopoietin-2 ELISA performs according to FDA/ICH/EMEA guidelines
Excellent correlation to existing methods
For affordable and efficient research
Interleukin-6 (IL-6) is a multifunctional cytokine that plays an important role in the body´s response to tissue injury or infection. Il-6 is produced by various cells, including T-cells, B-cells, macrophages, as well as vascular endothelial cells, mast cells, and dentritic cells (1). Il-6 secretion is stimulated during inflammatory response and travels through the bloodstream to the liver, where it induces the production of acute phase reactants such as C-reactive protein and others (1).
Human Interleukin-6 ELISA Assay Kit – high sensitivity
BIOMEDICA´s human Interleukin-6 (IL-6) ELISA Assay is highly sensitive and shows detectable values in serum and in plasma samples.
Features&Benefits
High Specificity – using characterized epitope-mapped antibodies
High Sensitivity – measurablevalues inbothserum and plasma
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Standardized – use of WHO intern.standards for kit calibration & harmonization
User-Friendly – ready to use color coded prediluted standards & controls
Take 15% off the human IL-6 ELISA Assay now through end of 2024!
Inflammatory diseases: elevated IL-6 levels are detected in various inflammatory conditions such as rheumatoid arthritis (2), systemic lupus erythematosus (3) , and inflammatory bowel disease (4). Therapies targeting Il-6 have been already approved (5).
Cancer: Il-6 is abundantly present in the tumor microenvironment of various tumours. It promotes tumnorigenesis, invasiveness, and metastases in cancer (6). New perspectives in cancer immunotherapy targeting IL-6 are discussed (7).
Metabolic diseases: Dysregulated IL-6 activity is associated with pathologies involving chronic inflammation and autoimmunity including metabolic diseases like obesity, metabolic syndrome or type-2 diabetes (8).
Kidney disease is estimated to affect more than 800 million people worldwide (1). Taking action for prevention, early diagnosis and treatment can reduce the risk and the burden of kidney disease.
Role of the kidneys, kidney disease and risk factors
Kidneys are vital organs that regulate fluid balance, blood pressure and produce hormones that stimulate the production of red blood cells . Kidney disease is a condition in which kidneys lose their ability to effectively filter waste products and excess fluids from the blood. Kidney disease commonly leads to a decline in kidney function that may lead to kidney failure, characterized by the complete loss of kidney function. At this stage dialysis or kidney transplantation become the only treatment option.
Kidney problems can emerge suddenly (acute) or gradually over time (chronic). Various conditions, diseases and medications can contribute to acute and chronic kidney problems. Chronic kidney disease (CKD) is characterized by a prolonged period of kidney abnormalities that last for more than three months (2), whereas acute kidney disease – acute kidney injury (AKI) is characterized by a sudden loss of excretory kidney function (3).
Other forms of kidney disease include polycystic kidney disease (PKD) a genetic disorder that leads to kidney enlargement and impaired kidney function over time and glomerulonephritis (GN). GN is a group of diseases characterized by inflammation of the glomeruli, the filtration units of the kidney (4).
The most common risk factors for the development and progression of CKD are diabetes and high blood pressure. Managing blood sugar and blood pressure can help keep kidneys healthy. Other risk factors of CKD include heart disease, obesity, family history and genetic background as well as age, smoking and nutrition (5).
Consequences of kidney disease include heart disease, high blood pressure, bone and mineral disorders, and anemia (6).
Keep Your Kidneys Healthy
Regular exercise, weight control, a balanced diet (7) and sufficient fluid intake are only some of the ways to keep your kidney healthy.
World Kidney Day – about kidney health download here and check out the 6-Step Guide to Protecting Kidney Health here .
BIOMEDICA – Biomarker ELISA kits for clinical research in kidney disease
HIGH QUALITY ASSAYS – Fully validated according to international quality guidelines
It is well recognized that exercise has beneficial effects on overall health (1). Skeletal muscle has been described as a “secretory organ” that produces cytokines in response to muscle contraction (2). Thus, muscle communicates with various organs and tissues e.g. adipose tissue, liver, pancreas, bones, and brain (1).
IL-6 in exercise and health
Interleukin-6 (IL-6) is a cytokine that is primarily known to play a key role in inflammation and the regulation of the immune response. However, IL-6 is also an important signaling molecule during exercise. Numerous studies have demonstrated that Il-6 is produced locally in working skeletal muscle (3, 4). Exercise, in particular intense or prolonged physical activity increases circulating IL-6 levels contributing to the regulation of glucose and lipid metabolism (4). Additional effects of IL-6 are discussed in Exercise and health — emerging roles of IL-6.
We at BIOMEDICA develop and manufacture ELISA assays to reliably quantify cytokines in biological fluids.
Our ELISA kits are rigorously validated according to international quality standards to ensure their consistency, specificity, precision, and robustness.
All of the validation data can be found on the respective product pages on our website www.bmgrp.com .
Quantify Cytokines with an ELISA Assay from BIOMEDICA
We designed our assays to quantify the cytokine of interest in serum, plasma or cell culture supernatants.
Standardized harmonization of results with WHO reference reagents
To ensure a harmonized standardization, WHO reference reagents / standards are employed for cross-comparison of results (1)
The IL-6 (Interleukin-6) ELISA kit from BIOMEDICA is a conventional sandwich ELISA kit. The assay incorporates two epitope-mapped antibodies that specifically bind to human IL-6 in the respective samples.
Capture antibody : is a recombinant IL-6 antibody that is specific for human IL-6, which is pre-coated on a 96-well microtiter plate.
Detection antibody: is a polyclonal IL-6 antibody (labeled with streptavidin-HRPO), which is specific for human IL-6.
Assay method: sandwich ELISA
Sample types: serum, plasma, cell-culture, urine
Sample volume: 100µl / well
Assay time: 4.5 hrs
Standard range: 0; 3.1 – 200 pg/ml
Sensitivity: LLOQ: 0.78 pg/ml (measurable concentrations in both serum AND plasma samples!), LOD: 0.28 pg/ml
The VEGF (Vascular Endothelial Growth Factor) ELISA kit from BIOMEDICA is a conventional sandwich ELISA kit. The assay incorporates two epitope-mapped antibodies that specifically bind to human VEGF in the respective samples.
Capture antibody is a recombinant VEGF antibody that is specific for human VEGF, which is pre-coated on a 96-well microtiter plate, recognizing a structural epitope in the conserved receptor binding- site of the VEGF peptide. All VEGF isoforms are detected.
Detection antibody: is a polyclonal VEGF antibody (labeled with streptavidin-HRPO), which is specific for human VEGF. The antibody recognizes multiple linear epitopes that are concentrated in the first 120 amino acids in the N-terminal region of the VEGF molecule.
The ANG2 (Angiopoietin-2) ELISA kit from BIOMEDICA is a conventional sandwich ELISA kit. The assay incorporates two epitope-mapped antibodies that specifically bind to human Angiopoietin-2 in the respective samples.
Capture antibody is a recombinant ANG2 antibody that is specific for human VEGF, which is pre-coated on a 96-well microtiter plate. The capture antibody has a structural epitope that binds to the receptor binding site of human Angiopoietin-2. The receptor binds to TEL / TIE2 protein.
Detection antibody: is a polyclonal ANG2 antibody (labeled with streptavidin-HRPO), which is specific for human ANG2. The antibody recognizes several linear epitopes that are distributed over the entire Angiopoietin-2 molecule.
Specificity: the ELISA recognizes endogenous (natural) human ANG2 and recombinant ANG2. Most probably all three ANG2 isoforms are detected by the assay, as determined by epitope mapping and analysis of the ANG2 sequence. This ELISA assay does not detect ANG1.
Standard matrix: 8 x vials of pre-diluted standards, lyophilized, in a serum based matrix containing recombinant human ANG2.
Controls: 2 controls (high and low) included in the kit
Mouse /rat Angiopoietin-2 ELISA (cat. no. BI-ANG2MR) also available
Major depressive disorder (MDD) is a prevalent mental health condition that ranks among the top psychiatric illnesses worldwide. Approximately 30-60% of patients diagnosed with depression do not show positive responses to currently available antidepressant treatments. There is consistent evidence suggesting that elevated blood levels of the cytokine interleukin-6 (IL-6) in individuals with MDD have a significant impact on stress responses and correlate with depression severity scores (1, 2).
IL-6 in depressive disorder
Given the association between IL-6 and MDD, inhibiting IL-6 has gained attention as a potential therapeutic approach for depression. By targeting IL-6 pathways it is hypothesized that the aberrant inflammatory response associated with MDD can be modulated, leading to improved treatment outcomes. Exploring IL-6 inhibition, as a novel strategy holds promise for the development of more effective therapies for individuals who are non-responsive to current antidepressant treatments.
IL-6 can reliably be measured in various sample types by ELISA assay:
Depression is common in women with obstructive sleep apnea (OSA), but objective markers of depression have not yet been explored in such patients. We hypothesized that inflammation and antioxidant biomarkers may be associated with depression in a cohort of OSA women. We conducted a multicentre, cross-sectional study in 247 women diagnosed with moderate-to-severe OSA. Depression was assessed by the depression subscale of the Hospital Anxiety and Depression Questionnaire (HAD-D) and defined as a score ≥11. Associations between tumour necrosis factor α (TNFα), interleukin 6 (IL-6), C-reactive protein (CRP), intercellular adhesion molecule 1 (ICAM-1), catalase (CAT), superoxide dismutase (SOD) and brain-derived neurotrophic factor (BDNF) plasma levels and depression were assessed. The women had a median (25th-75th percentiles) age of 58 (51-65) years, body mass index (BMI) of 33.5 (29.0-38.3) Kg/m2 , Epworth Sleepiness Scale (ESS) score of 10 (6-13) and apnea-hypopnea index (AHI) of 33.3 (22.8-49.3). Logistic regression analyses revealed that only IL6 levels were associated with the presence of depression (adjusted odds ratio [OR], 1.20; 95% confidence interval [CI], 1.08-1.34), whereas linear regression further confirmed that IL6 levels were significantly associated with HAD-D scores (β = .154; 95% CI, 0.03-0.30). Multivariate regression analysis showed that IL6 (OR, 1.22; 95% CI, 1.09-1.36), ESS (OR, 1.10; 95% CI 1.02-1.19) and physical activity <30 min/day (OR, 2.51; 95% CI, 1.25-5.05) were independent predictors of depression. Thus, we conclude that in a cohort of women with moderate-to-severe OSA, IL6 levels are independently associated with the presence of depression and correlate with depression scores. Low physical activity and higher ESS scores are also independent indicators of risk of depression in this population.
At Biomedica we develop and manufacture quality ELISAs to quantify cytokine release
Our ELISAs to quantify cytokine release are validated according to international quality guidelines to ensure their consistency, specificity, precision, and robustness. The validation data files can be found on the respective product pages www.bmgrp.com.
Our kits are designed to specifically quantify cytokine release in various biological matrices (e.g. serum, plasma, cell culture).
WHO reference reagents for a harmonized standardization
All our cytokine kits are standardized using WHO reference reagents/standards to allow cross-comparison of results when using different reagent sets during a study (1).
The Biomedica IL-6 (interleukin-6) ELISA kit is a sandwich ELISA that incorporates two epitope-mapped antibodies that specifically bind to human IL-6 in the respective samples.
Capture antibody (pre-coated on a 96-well microtiter plate): recombinant IL-6 antibody (specific for human IL-6).
Detection antibody: polyclonal IL-6 antibody (streptavidin-HRPO labeled), specific for human IL-6.
Method: sandwich ELISA
Sample type: serum, plasma, cell-culture, urine
Sample volume: 100µl / well
Assay time: 4.5 hrs
Standard range: 0 / 3.1 – 200 pg/ml
Sensitivity: LOD: 0.28 pg/ml; LLOQ: 0.78 pg/ml (measurable concentrations in serum AND plasma samples!)
The Biomedica VEGF (Vascular Endothelial Growth Factor) ELISA kit is a sandwich ELISA that incorporates two epitope-mapped antibodies that specifically bind to human VEGF in the respective samples.
Capture antibody (pre-coated on a 96-well microtiter plate): recombinant VEGF antibody (specific for human VEGF) recognizes a structural epitope in the conserved receptor binding- site of the VEGF peptide. The antibody binds to all VEGF isoforms.
Detection antibody: polyclonal VEGF antibody (streptavidin-HRPO labeled), specific for human VEGF, recognizing multiple linear epitopes that are concentrated in the first 120 amino acids in the N-terminal region of the VEGF molecule.
Method: sandwich ELISA
Sample type: serum, plasma, cell-culture, urine
Sample volume: 10µl / well
Assay time: 4.5 hrs
Standard range: 0 / 31.2 – 2000 pg/ml
Sensitivity: LOD: 2.5 pg/ml; LLOQ: 15.6 pg/ml (measurable concentrations in serum AND plasma samples)
The Biomedica ANG2 (Angiopoietin-2) ELISA kit is a sandwich ELISA that incorporates two epitope-mapped antibodies that specifically bind to human Angiopoietin-2 in the respective samples.
Capture antibody (pre-coated on a 96-well microtiter plate): recombinant ANG2 antibody (specific for human ANG2).The capture antibody has a structural epitope that binds to the receptor binding site of human Angiopoietin-2. The receptor binds to TEL / TIE2 protein.
Detection antibody: polyclonal ANG2 antibody (streptavidin-HRPO labeled), specific for human ANG2, recognizing several linear epitopes that are distributed over the entire Angiopoietin-2 molecule.
Specificity: the ELISA recognizes recombinant and endogenous (natural) human ANG2. The assay most probably detects all three ANG2 isoforms, as determined by epitope mapping and analysis of the ANG2 sequence. ANG1 is not detected in this ELISA assay.
Standard matrix: Serum based matrix containing recombinant ANG2, including 8x vials of pre-diluted standards, lyophilized
Controls: 2 controls (high and low) included
Also available: Mouse /rat Angiopoietin-2 ELISA (cat. no. BI-ANG2MR)
Interleukin-6 (IL-6) is a key immunomodulatory cytokine that affects the pathogenesis of diverse diseases, including autoimmune diseases, chronic inflammatory conditions and cancer. Classical IL-6 signalling involves the binding of IL-6 to the membrane-bound IL-6 receptor α-subunit (hereafter termed ‘mIL-6R’) and glycoprotein 130 (gp130) signal-transducing subunit. By contrast, in IL-6 trans-signalling, complexes of IL-6 and the soluble form of IL-6 receptor (sIL-6R) signal via membrane-bound gp130. A third mode of IL-6 signalling – known as cluster signalling – involves preformed complexes of membrane-bound IL-6-mIL-6R on one cell activating gp130 subunits on target cells. Antibodies and small molecules have been developed that block all three forms of IL-6 signalling, but in the past decade, IL-6 trans-signalling has emerged as the predominant pathway by which IL-6 promotes disease pathogenesis. The first selective inhibitor of IL-6 trans-signalling, sgp130, has shown therapeutic potential in various preclinical models of disease and olamkicept, a sgp130Fc variant, had promising results in phase II clinical studies for inflammatory bowel disease. Technological developments have already led to next-generation sgp130 variants with increased affinity and selectivity towards IL-6 trans-signalling, along with indirect strategies to block IL-6 trans-signalling. Here, we summarize our current understanding of the biological outcomes of IL-6-mediated signalling and the potential for targeting this pathway in the clinic.
IL-6 (cat. no. BI-IL6) – high sensitivity – detectable levels
VEGF (cat. no. BI-VEGF) – low sample volume – 10 µl – detecting all known VEGF isoforms
ANGIOPOIETIN-2 (cat. no. BI-ANG2) / optimized assay range
Also available mouse/rat Angiopoietin-2 ELISA kit (cat. no. BI-ANG2MR) – only 5µl sample volume.
Biomedica ELISA kit – including color coded reagents
ABOUT CYTOKINES
Cytokines are critical cell signaling molecules that play important roles in the immune response and many other physiological processes. These proteins are produced by a wide variety of cell types that act on specific target cell receptors to trigger a range of biological responses.
Cytokines are involved in many aspects of the immune response, including inflammation, cell growth and differentiation, and cell death (apoptosis). Cytokines also regulate tissue repair and wound healing. Dysregulation of cytokine production and signaling may contribute to various diseases including autoimmune disorders, allergies, and cancer.
Some common cytokines include interleukins, tumor necrosis factor (TNF), and interferons.
Interleukin-6 (IL-6) is a pleiotropic cytokine that is largely involved in immunomodulatory processes. One of its roles is to respond to infections. As a pro-inflammatory cytokine, IL-6 activates the immune response, recruiting immune cells triggering B and T cell response. IL-6 dysregulation is associated with pathologies involving chronic inflammation, autoimmune diseases, and atherosclerosis. However, IL-6 also has beneficial anti-inflammatory and positive metabolic effects that are released during exercise coupled to a physical adaptation to intense training. In addition to its immune functions, IL-6 is involved in many other physiological processes, including bone remodeling and the nervous system.
Angiopoietin-2 (ANG2) is a protein that plays a critical role in regulating the development and remodeling of blood vessels, a process known as angiogenesis. ANG-2 is primarily produced by endothelial cells, which are cells that line the interior of blood vessels. Under normal physiological conditions the expression of ANG2 is low, which allows the blood vessels to undergo a rapid growth and remodeling. However, in pathological conditions, such as inflammation and cancer, ANG2 expression can be upregulated, leading to excessive angiogenesis and blood vessel destabilization.
Vascular endothelial growth factor (VEGF) is a protein mainly produced by endothelial cells. VEGF stimulates the growth and proliferation of endothelial cells, as well as the migration and differentiation of these cells into new blood vessels. VEGF plays a role in the development of cancer and certain ocular diseases, where it promotes the growth of new blood vessels that feed tumors or damage the retina.
There are several different isoforms of VEGF, each with different properties and functions. For example, VEGF-A is the most well-known cytokine of the VEGF family, and is primarily responsible for promoting angiogenesis. Other isoforms, such as VEGF-B and VEGF-C, have different roles in the body, such as regulating the growth of blood vessels in the heart and lymphatic system, respectively.
Bone metastases affect over 1.5 million cancer patients globally, making bones a favored metastatic site for solid tumors (1). The presence of skeletal metastases can significantly reduce the quality of life for individuals with advanced cancer, as weakened bones can cause pain, fractures, and increase the risk of mortality (2, 3).
Bones have numerous important functions which include structural support, mobility, hematopoiesis, and mineral storage (4). The health and performance of our bone tissue are overseen by a range of cell types, which include:
Osteoblasts – cells responsible for creating new bone tissue
Osteoclasts – cells responsible for breaking down bone tissue
Osteocytes – cells within the bone that monitor mechanical loading and regulate the process of bone remodeling
Bone biology- cancer and bone
Cancer and Bone – bone a preferred target site for cancer metastasis
Bone is a favored destination for metastasis in specific types of cancer, whereby cancer cells detach from the primary tumor and disseminate throughout the body. Certain cancers, such as breast, prostate, kidney, lung, ovarian, and thyroid, are especially prone to spread to bone (5).
Cancer and Bone – the RANK/RANKL/OPG system
The RANK/RANKL/OPG system (receptor activator of the nuclear factor-κB ligand/ Osteoprotegerin) was identified more than 20 years ago and remains a widely researched topic until this day. The interaction between RANK and RANKL is essential for bone metabolism and osteoclast development. OPG acts as a decoy receptor for RANKL. By binding completely to RANKL, OPG obstructs the RANKL-RANK interaction, thereby blocking bone resorption. In addition to its role in regulating bone remodeling, the RANK/RANKL/OPG system is directly implicated in tumor cell development, particularly in the progression of breast and prostate cancer as well as leukemia (6-8).
Inhibiting RANK/RANKL signaling in human cancer
A fully humanized monoclonal antibody has been developed to counteract the effects of RANKL, which has received approval for treating bone loss conditions. The antibody functions by disrupting RANK signaling, preventing osteoclast activation and inhibiting bone resorption (6-8). More recently, inhibiting RANKL has been recognized as a significant checkpoint with the potential to influence anti-tumor response. Consequently, RANKL blockade has a direct impact on bone metastasis. Ongoing clinical and experimental trials are evaluating this emerging therapeutic approach (9).
Studies analyzing serum levels of OPG and RANKL
Serum concentrations of both OPG and soluble RANKL can be accessed via an enzyme-linked immunosorbent assay (ELISA). The predictive value of RANKL/OPG serum levels and disseminated tumor cells in breast cancer patients without metastasis was investigated in 509 patients with primary, nonmetastatic breast cancer. The results demonstrated that RANKL serum levels were significantly elevated in patients who developed bone metastases (10). A different study highlighted the role of OPG as a marker of breast cancer risk in women with a BRCA1 mutation. The authors suggested that OPG levels could be associated with disease risk, potentially serving as a marker for breast cancer risk and improving existing risk prediction models by identifying women at high risk of developing the disease (11).
How are circulating serum levels of Osteoprotegerin and soluble RANKL measured?
Both markers can easily be measured with a conventional ELISA assay.
Understanding the Bone in Cancer Metastasis. Fornetti J, Welm AL, Stewart SA. J Bone Miner Res. 2018 Dec;33(12):2099-2113. doi: 10.1002/jbmr.3618. Epub 2018 Nov 26. PMID: 30476357.
Cancer to bone: a fatal attraction. Weilbaecher KN, Guise TA, McCauley LK. Nat Rev Cancer. 2011 Jun;11(6):411-25. doi: 10.1038/nrc3055. Epub 2011 May 19. PMID: 21593787; PMCID: PMC3666847.
Bone metastasis: mechanisms, therapies, and biomarkers. Clézardin P, Coleman R, Puppo M, Ottewell P, Bonnelye E, Paycha F, Confavreux CB, Holen I. Physiol Rev. 2021 Jul 1;101(3):797-855. doi: 10.1152/ physrev.00012.2019. Epub 2020 Dec 24. PMID: 33356915.
The Roadmap of RANKL/RANK Pathway in Cancer. Casimiro S, Vilhais G, Gomes I, Costa L. Cells. 2021 Aug 4;10(8):1978. doi: 10.3390/cells10081978. PMID: 34440747; PMCID: PMC8393235.
RANKL and RANK in Cancer Therapy. Physiology (Bethesda). Onji M, Penninger JM. 2023 May 1;38(3):0. doi: 10.1152/physiol.00020.2022. Epub 2022 Dec 6. PMID: 36473204.
World Kidney Day – March 9, 2023 – raising awareness of the importance of our kidneys
Around 1 in 10 people suffer from chronic kidney disease (CKD), with more than 800 million individuals being affected worldwide. CKD is a progressive disease in which the kidneys gradually lose their function over time. If detected early, medication and changes in lifestyle may help to prevent or slow down CKD progression.
What are the risk factors for CKD?
Older age (+60 years), diabetes, high blood pressure, heart disease, obesity and some medications are some known risk factors for kidney disease.
How can we keep our kidneys healthy?
Nutrition, exercise, sufficient fluid intake are only some examples on how to keep our kidneys healthy. Valuable information can be found on the following websites :
Purpose of review: Plant-based diets have been used with growing popularity for the treatment of a wide range of lifestyle-related diseases, including diabetes, hypertension, and obesity. With the reinvigoration of the dietary management of chronic kidney disease (CKD) and the use of low protein diets for secondary prevention of CKD to delay or prevent dialysis therapy, there is an increasing interest in the potential role of plant-based diets for these patients.
Recent findings: Recently, a body of evidence related to the role of plant-based diets in preventing CKD has reemerged. Several observational studies have shown that red and processed meat have been associated with increased risk of CKD as well as faster progression in those with preexisting CKD. In several substitution analyses, replacement of one serving of red and/or processed meat has been linked with sizable reductions in CKD risk. Although limited, experimental trials for the treatment of metabolic acidosis in CKD with fruits and vegetables show outcomes comparable to oral bicarbonate. The use of plant-based diets in CKD may have other benefits in the areas of hypertension, weight, hyperphosphatemia, reductions in hyperfiltration, and, possibly, mortality. The risk of potassium overload from plant-based diets appears overstated, mostly opinion-based, and not supported by the evidence. Plant-based diets are generally well tolerated and provide adequate protein intake, including essential amino acids as long as the diet is correctly implemented.
Summary: Plant-based diets should be recommended for both primary and secondary prevention of CKD. Concerns of hyperkalemia and protein inadequacy related to plant-based diets may be outdated and unsupported by the current body of literature. Healthcare providers in general medicine and nephrology can consider plant-based diets as an important tool for prevention and management of CKD.
Chronic kidney disease (CKD) is a major public health problem worldwide. Its prevalence and incidence are increasing, particularly among the ethnic minority populations. Diabetes, hypertension and obesity have been the three major aetiologies for CKD in all developed countries. While diabetes and hypertension remain the major causes of CKD in developing countries, environmental pollution, pesticides, water, analgesic abuse and herbal medications are common causes in these regions. Rapid urbanization and globalization are thought to be the contributing factors to rising prevalence and incidents of CKD. Despite the rising prevalence of CKD, disease awareness remains profoundly low. Worldwide, only 6% of the general population and 10% of the high-risk population are aware of their CKD statuses. Health screenings have been shown to be effective in improving the incidence of ESRD. However, currently there is no effective tool to assess and evaluate the awareness objectively.
Biomarkers & Bone Health – ELISA Kits for Clinical Research
Bone remodeling is a continuous process that removes bone and replaces it with newly synthesized bone. This bone turnover process preserves the mechanical function of the human skeleton.
Bone turnover biomarkers, e.g. markers of bone formation and bone resorption, have been used during the last decade to monitor bone diseases and to monitor their treatment.
Many of these markers are secreted by osteoblasts and osteoclasts and include regulators of bone turnover e.g. receptor activator of NF-kB ligand (RANKL) and osteoprotegerin (OPG).
Though RANKL and OPG play an integral role in bone turnover, they do not reflect the activity of osteocytes, the most abundant cell type in the bone.
Osteocytes are cells that regulate bone remodeling. They secrete proteins – bone regulation markers – that include Sclerostin (SOST), Dickkopf-1 (DKK-1), and Fibroblast growth factor (FGF23). These markers reflect the osteocyte activity.
The above listed biomarkers circulate and can be measured in serum and plasma allowing the investigation of complex interactions between the bone and their relationship with other organs.
+ EASY – ready to use calibrators & controls included (color-coded reagents) +FULL VALIDATION PACKAGE – assays are optimized for clinical samples + HIGH QUALITY GUARANTEED – results you can rely on + WIDELY CITED in 1500 + publications
Biomedica – Complete ready-to-use ELISA kits for superior performance and reproducibility
Biochemical markers of bone turnover have been used for decades in the management of bone diseases, to assess the prognosis of these conditions and to monitor treatments. The new markers, however, also reflect specific physiological mechanisms in the bone or other organs. Periostin may be more specific to the periosteum; cathepsin K is an osteoclastic enzyme that may be involved in the cardiovascular system and joints; Dickkopf-1 is involved in bone formation and vascular calcification; sclerostin is a major regulator of bone formation in response to mechanical loading and may also play a role in chronic kidney disease bone and mineral disorder; sphingosine-1-phosphate is a lipid mediator interacting with bone resorption. Some of the bone markers are in fact hormones produced by the bone that affect various physiological and pathological functions in other organs. Thus, osteocalcin is produced by osteoblasts and participates in the regulation of insulin sensitivity and fertility in men. Fibroblast growth factor 23 is produced by osteocytes to regulate phosphorus and 1,25(OH)2D3, but it also plays a major role in the adverse consequences of declining renal function, in particular with respect to the myocardium. Micro RNAs are single-stranded RNAs that regulate several pathways, including the development timing, organogenesis, cell apoptosis, proliferation and differentiation. Their serum concentration may reflect the links between bone physiology and certain conditions in other organs, for example, the cardiovascular system.
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INTRODUCTION: Angiopoietin-2 (Ang-2) is an important regulator of the angiopoietin-1/Tie-2 receptor signaling system on endothelial cells during angiogenesis. Disruption of this signaling leads to the loss of endothelial integrity and renders the endothelium response towards a variety of pro-inflammatory cytokines and growth factors. Thus, Ang-2 might lead to vascular micro-inflammation in patients with CKD (chronic kidney disease). Ang-2 levels increase with CKD stage, are associated with fluid overload and abnormal cardiac structure and predict mortality in patients with CKD stages 4–5. Although Ang-2 levels return toward normal after successful kidney transplantation, Ang-2 remains a putative cardiovascular risk factor in this population.
METHODS: An enzyme-linked immunosorbent assay for the detection of all three angiopoietin-2 isoforms in human serum and plasma was developed. Two high quality antibodies are combined in a sandwich test format: As capturing antibody a recombinant monoclonal antibody is used. A biotin-labeled polyclonal affinity-purified antibody serves for detection of the analyte. High resolution epitope mapping of the antibodies via peptide microarray technology allowed the identification of linear antibody epitopes. Technical performance and accuracy of the assay were assessed according to ICH/EMEA guidelines.
RESULTS: Microarray data illustrate the binding of the polyclonal detection antibody to human angiopoietin-2 spotted on a chip. Altogether, seven linear epitopes located N-terminal/at the center of angiopoietin-2 were detected. Most relevant linear epitopes are epitope e2 in the super clustering region and e6 near the fibrinogen-like domain, displaying a twofold higher fluorescent signal than the remaining epitopes. For the recombinant monoclonal antibody no fluorescence was recorded. This antibody recognizes a structural epitope within the C-terminus of angiopoietin-2, which covers the bioactive receptor-binding site of the protein. We expect to detect all described angiopoietin-2 isoforms, as the receptor-binding site is conserved and the majority of the polyclonal antibody epitopes are present in all isoforms. This assay is in conformance with ICH/EMEA/FDA guidelines. Validation data demonstrate its applicability in nephrological disorders including chronic kidney disease. Here we compare an apparently healthy population to chronic kidney disease samples on haemodialysis and kidney transplant samples.
CONCLUSIONS: This new angiopoietin-2 ELISA enables a quick and accurate quantification of all human angiopoietin-2 isoforms that are bioactive in chronic kidney disease and other nephrological disorders.
RELATED LITERATURE
Molnar et al. (2014): Circulating angiopoietin-2 levels predict mortality in kidney transplant recipients: a 4-year prospective case–cohort study. Transplant International, 27 (6): 541-52. 2. David et al. (2010): Circulating angiopoietin-2 levels increase with progress of chronic kidney disease. Nephrol. Dial. Transplant, 25: 2571-2579,
Tsai et al. (2017): The interaction between fluid status and angiopoietin-2 in adverse renal outcomes of chronic kidney disease. PLoS One, 12 (3): e173906.
Tsai et al. (2016): Angiopoietin-2, Angiopoietin-1 and subclinical cardiovascular disease in Chronic Kidney Disease. Scientific Reports, 6:39400
Osteoporosis Awareness –
Worldwide around 200 million women have osteoporosis. Osteoporosis is a disease that weakens bone. Bones become fragile and porous. From the outside, the osteoporotic bone is shaped like normal bone. However, the inside of the bone loses density and becomes weak. The risk of fractures become greater with age, due to the loss of minerals like calcium and phosphate. Osteoporosis most often affects bones in the hip, the spine and the wrist. Factors that influence bone health are nutrition, exercise and hormonal factors. Osteoporosis cannot be cured but delayed through early intervention and biomarker research has identified proteins that may help to identify patients at risk.
Osteoporosis prevention includes a balanced diet, containing sufficient amounts of calcium and vitamin D and regular exercise.
Exercises to protect or reduce your chance of fracture include regular weight-bearing exercise (eg. weight lifting or resistance training) or any kind of activity that carries your own body weight (e.g. walking, running, climbing stairs, dancing).
A healthy and balanced diet with the recommended daily amounts of calcium are important for bone health. Read more about “Good for your bone foods .
Related literature
The Effectiveness of Physical Exercise on Bone Density in Osteoporotic Patients.
Benedetti MG, Furlini G, Zati A, Letizia Mauro G. Biomed Res Int. 2018. 23;2018:4840531. PMID: 30671455. Link to full text
Abstract
Physical exercise is considered an effective means to stimulate bone osteogenesis in osteoporotic patients. The authors reviewed the current literature to define the most appropriate features of exercise for increasing bone density in osteoporotic patients. Two types emerged: (1) weight-bearing aerobic exercises, i.e., walking, stair climbing, jogging, and Tai Chi. Walking alone did not appear to improve bone mass; however it is able to limit its progressive loss. In fact, in order for the weight-bearing exercises to be effective, they must reach the mechanical intensity useful to determine an important ground reaction force. (2) Strength and resistance exercises: these are carried out with loading (lifting weights) or without (swimming, cycling). For this type of exercise to be effective a joint reaction force superior to common daily activity with sensitive muscle strengthening must be determined. These exercises appear extremely site-specific, able to increase muscle mass and BMD only in the stimulated body regions. Other suggested protocols are multicomponent exercises and whole body vibration. Multicomponent exercises consist of a combination of different methods (aerobics, strengthening, progressive resistance, balancing, and dancing) aimed at increasing or preserving bone mass. These exercises seem particularly indicated in deteriorating elderly patients, often not able to perform exercises of pure reinforcement. However, for these protocols to be effective they must always contain a proportion of strengthening and resistance exercises. Given the variability of the protocols and outcome measures, the results of these methods are difficult to quantify. Training with whole body vibration (WBV): these exercises are performed with dedicated devices, and while it seems they have effect on enhancing muscle strength, controversial findings on improvement of BMD were reported. WBV seems to provide good results, especially in improving balance and reducing the risk of falling; in this, WBV appears more efficient than simply walking. Nevertheless, contraindications typical of senility should be taken into account.
Exercise for the prevention of osteoporosis in postmenopausal women: an evidence-based guide to the optimal prescription.
Daly RM, Dalla Via J, Duckham RL, Fraser SF, Helge EW.Braz J Phys Ther. 2019.23(2):170-180. PMID: 30503353. Full text link
Abstract
Background: Osteoporosis and related fragility fractures are a global public health problem in which pharmaceutical agents targeting bone mineral density (BMD) are the first line of treatment. However, pharmaceuticals have no effect on improving other key fracture risk factors, including low muscle strength, power and functional capacity, all of which are associated with an increased risk for falls and fracture, independent of BMD. Targeted exercise training is the only strategy that can simultaneously improve multiple skeletal and fall-related risk factors, but it must be appropriately prescribed and tailored to the desired outcome(s) and the specified target group.
Objectives: In this review, we provide an overview of the general principles of training and specific loading characteristics underlying current exercise guidelines for the prevention of osteoporosis, and an update on the latest scientific evidence with regard to the type and dose of exercise shown to positively influence bone mass, structure and strength and reduce fracture risk in postmenopausal women.
Breast Cancer – the most common cancer worldwide
Breast cancer (BC) is the most commonly occurring cancer in women and the most common cancer overall. Although it is mostly found in women, it can affect men as well. Breast tissue in men can also become malignant. Though male BCs are rare and occur in 1% of all BCs, men are often diagnosed at a more advanced stage. The delay in seeking medical attention often results in late presentation and poor prognosis.
October is breast cancer month – raising global awareness on risks, the importance of screenings, and the options of treatment.
Wilkinson L et al., Br J Radiol. 2022. PMID: 34905391; PMCID. Full text
Abstract
Breast cancer is now the most commonly diagnosed cancer in the world. The most recent global cancer burden figures estimate that there were 2.26 million incident cases in 2020 and the disease is the leading cause of cancer mortality in women worldwide. The incidence is strongly correlated with human development, with a large rise in cases anticipated in regions of the world that are currently undergoing economic transformation. Survival, however, is far less favourable in less developed regions. There are a multitude of factors behind disparities in the global survival rates, including delays in diagnosis and lack of access to effective treatment. The World Health Organization’s new Global Breast Cancer Initiative was launched this year to address this urgent global health challenge. It aims to improve survival across the world through three pillars: health promotion, timely diagnosis, and comprehensive treatment and supportive care.
regulator of tumor biology – expressed by endothelial cells – isoform-specific receptors for VEGF – checkpoint target – association with poor prognosis in BC patients.
extracellular matrix protein – novel therapeutic target – marker of glioma malignancy and potential tumor recurrence – high serum levels associated with a poor survival in BC patients.
Leucine-rich alpha-2-glycoprotein 1 – LRG1 – involved in pathogenic angiogenesis in cancer – wide spread in the microenvironment of numerous tumors – contributes to vascular dysfunction – potential therapeutic target.
important cytokine during breast cancer progression – IL6 triggers activation of STAT2 in breast tumors – soluble factor IL6 could be used for early diagnosis of BC or prevent development of metastasis to the bone.
Cancer research for the prevention and early recognition of cancer
Promoting cancer research to beat cancer: prevention and early detection of cancer is essential to control the disease. Understanding the causes of cancer and the ability to detect cancer sooner has a great impact on the survival and the outcome of patients.
Subsequently, early detection strategies and progress in diagnostic procedures will help to develop treatments to control the disease.
Promoting cancer research:
Exploring new biomarkers may help to identify the disease early.
“Early diagnosis of cancer focuses on detecting symptomatic patients as early as possible so they have the best chance for successful treatment. When cancer care is delayed or inaccessible there is a lower chance of survival, greater problems associated with treatment and higher costs of care. Early diagnosis improves cancer outcomes by providing care at the earliest possible stage and is therefore an important public health strategy in all settings…” Read more
Pelosi E, Castelli G, Testa U.Ann Ist Super Sanita. 2019 Oct-Dec;55(4):371-379. doi: 10.4415/ANN_19_04_11. PMID: 31850865.
Abstract
The burden of cancer is increasing worldwide, with a continuous rise of the annual total cases. Although mortality rates due to cancer are declining in developed countries, the total number of cancer deaths continues to rise due to the increase in the number of aged people. Three main causes of cancer have been described, represented by environmental factors, hereditary factors and random factors related to defects originated during cell replication. The frequency of cancers is very different for the various tissues and there is great debate on the extent of the specific contribution of environmental factors and random factors (due to “bad luck”) to cancer development. However, there is consensus that about 50% of all cases of cancer are related to environment and are preventable. Although a part of cancers is related to intrinsic mechanisms non preventable of genetic instability, it is evident that implementation of primary and secondary prevention measures is the only affordable strategy to meet from a medical and economic point of view the tremendous pressure created on healthcare structures by the increased cancer burden. It is time to bypass the paradox of disease prevention: celebrated in principle, resisted in practice.
Ovarian cancer is a silent cancer which rate survival mainly relays in early stage detection. The discovery of reliable ovarian cancer biomarkers plays a crucial role in the disease management and strongly impact in patient’s prognosis and survival. Although having many limitations CA125 is a classical ovarian cancer biomarker, but current research using proteomic or metabolomic methodologies struggles to find alternative biomarkers, using non-invasive our relatively non-invasive sources such as urine, serum, plasma, tissue, ascites or exosomes. Metabolism and metabolites are key players in cancer biology and its importance in biomarkers discovery cannot be neglected. In this chapter we overview the state of art and the challenges facing the use and discovery of biomarkers and focus on ovarian cancer early detection.
Keywords: Cancer biomarkers; Early detection; Metabolomics; Ovarian cancer; Proteomics; Urine biomarkers.
Developed & manufactured by Biomedica – Biomarkers for Cancer Research
The identification and validation of biomarkers in cancer is essential to improve our understanding of the disease. The emergence of novel cancer biomarkers continues to grow as scientists strive to find promising novel therapeutic targets and new prognostic and predictive markers to fight the disease.
• RELIABLE – full validation package
• CONVENIENT – assay range optimized for clinical samples
• EASY – ready to use prediluted calibrators & controls
• LOW sample volumes
• TRUSTED – widely cited
High specificity – known target binding sites through mapping data
The unique specificity of the proprietary antibodies used in the Biomedica ELISA kits ensure that the assays only measure the analyte of interest.
Biomarkers for Cancer Research – Learn more about the markers
The transmembrane protein Neuropilin-1 (NRP1) regulates tumor biology and has been identified as a checkpoint target (1). High tissue NRP-1 levels are associated with a poor prognosis in breast cancer patients. In a recent study (2), German researchers have shown that circulating soluble NRP1 serum levels are an independent marker for poor prognosis in early breast cancer. Soluble Neuropilin-1 was quantified in serum with the highly specific NRP1 ELISA from Biomedica. Therapeutic areas of NRP1.
The secreted extracellular matrix protein Periostin has evolved as a novel therapeutic target and is a robust marker of glioma malignancy and potential tumor recurrence. It has also been implicated in the pathogenesis of breast cancer as high serum levels of periostin are associated with a poor survival in breast cancer patients (3). Periostin was quantified in serum with the well characterized Biomedica Periostin ELISA that has been published (4). Therapeutic areas of Periostin.
Semaphorin 4D (Sema4D) is a glycoprotein that is emerging as clinical biomarker and as therapeutic target in cancer. It has been associated with cancer progression and the occurrence of bone metastases (5, 6). Therapeutic areas of Sema4D.
Leucine-rich alpha-2-glycoprotein 1 (LRG1) is a protein that is an important factor involved in pathogenic angiogenesis in cancer. It is abundantly present in the microenvironment of many tumors contributing to vascular dysfunction and thus serving as a potential therapeutic target (7). Therapeutic areas of LRG 1.
The RANKL/RANK/OPG system contributes to the development of bone metastases and influences tumor biology in earlier stages of cancer (9). Dysregulation has been widely documented in the context of metastatic bone disease (10). The Biomedica OPG and RANKL ELISA kits have been widely used in the respective studies.
Literature
1. Neuropilin-1: a checkpoint target with unique implications for cancer immunology and immunotherapy. Chuckran CA et al., J Immunother Cancer. 2020. 8(2):e000967.
2. Soluble Neuropilin-1 is an independent marker of poor prognosis in early breast cancer. Rachner TD et al., J Cancer Res Clin Oncol. 2021. 147(8):2233-2238.
3. High serum levels of periostin are associated with a poor survival in breast cancer. Rachner TD et al., 2020. 180(2):515-524.
4. Characterization of a sandwich ELISA for the quantification of all human periostin isoforms. Gadermaier E J Clin Lab Anal. 2018. 32(2):e22252.
5. Plasma levels of Semaphorin 4D are decreased by adjuvant tamoxifen but not aromatase inhibitor therapy in breast cancer patients. Göbel A J Bone Oncol. 2019. 4;16:100237.
6. Semaphorins as emerging clinical biomarkers and therapeutic targets in cancer. Mastrantonio R et al., Theranostics. 202. 15;11(7):3262-3277.
7. Leucine-rich alpha-2-glycoprotein 1 (LRG1) as a novel ADC target. Javaid F et al., RSC Chem Biol. 2021. 31;2(4):1206-1220.
8. RANKL/RANK/OPG system beyond bone remodeling: involvement in breast cancer and clinical perspectives. Infante M J Exp Clin Cancer Res. 2019. 8;38(1):12.
9. Serum receptor activator of nuclear factor κB ligand (RANKL) levels predict biochemical recurrence in patients undergoing radical prostatectomy. Todenhöfer T, BJU Int. 2014. 113(1):152-9.
10. Prognostic Value of RANKL/OPG Serum Levels and Disseminated Tumor Cells in Nonmetastatic Breast Cancer. Rachner TD et al., Clin Cancer Res. 2019. 15;25(4):1369-1378.
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Related information:
Our Biomedica human Angiopoietin-2 ELISA (cat.no. BI-ANG2) has been cited in the following publication:
Testicular Cancer – RANKL inhibition suppresses tumor growth: Testicular cancer is the most commonly diagnosed malignancy in young males. A new study features the use of the Biomedica RANKL ELISA investigating the effects of RANKL (receptor activator of nuclear factor- κB ligand) inhibition in patients with testicular cancer. Despite its high cure rate, patients suffer from serious adverse effects to chemotherapy. Investigating alternative treatments, the scientists demonstrated that the RANKL inhibitor Denosumamb, used to treat osteoporosis, has tumor suppressive properties. As RANKL signaling was recently identified in the testis regulating male reproductive function, an open question remains if RANKL is responsible for the malignant transformation to invasive tumors.
Testicular Cancer – RANKL inhibition suppresses tumor growth
Andreassen CH, Lorenzen M, Nielsen JE, Kafai Yahyavi S, Toft BG, Ingerslev LR, Clemmensen C, Rasmussen LJ, Bokemeyer C, Juul A, Jørgensen A, Blomberg Jensen M. Br J Cancer. 2022 Apr 13. doi: 10.1038/s41416-022-01810-w. Epub ahead of print. PMID: 35418213.
Abstract
Background: Testicular germ cell tumours (TGCTs) have a high sensitivity to chemotherapy and a high cure rate, although with serious adverse effects. In the search for tumour suppressive drugs, the RANKL inhibitor Denosumab, used to treat osteoporosis, came up as a candidate since RANKL signalling was recently identified in the testis.
Methods: Expression of RANKL, RANK and OPG, and the effects of RANKL inhibition were investigated in human TGCTs, TGCT-derived cell-lines, and TGCT-xenograft models. Serum RANKL was measured in TGCT-patients.
Results: RANKL, RANK, and OPG were expressed in germ cell neoplasia in situ (GCNIS), TGCTs, and TGCT-derived cell lines. RANKL-inhibition reduced proliferation of seminoma-derived TCam-2 cells, but had no effect on embryonal carcinoma-derived NTera2 cells. Pretreatment with Denosumab did not augment the effect of cisplatin in vitro. However, inhibition of RANKL in vivo reduced tumour growth exclusively in the TCam-2-xenograft model and Denosumab-treatment decreased proliferation in human GCNIS cultures. In TGCT-patients serum RANKL had no prognostic value.
Conclusions: This study shows that the RANKL signalling system is expressed in GCNIS and seminoma where RANKL inhibition suppresses tumour growth in vitro and in vivo. Future studies are needed to determine whether RANKL is important for the malignant transformation or transition from GCNIS to invasive tumours.
Widely cited in clinical studies – 290+ publications
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Blomberg Jensen M, Andreassen CH, Jørgensen A, Nielsen JE, Juel Mortensen L, Boisen IM, et al. Nat Commun. 2021;12:1–15. doi: 10.1038/s41467-021-22734-8. PMID: 33893301; PMCID: PMC8065035.
Abstract
Infertile men have few treatment options. Here, we demonstrate that the transmembrane receptor activator of NF-kB ligand (RANKL) signaling system is active in mouse and human testis. RANKL is highly expressed in Sertoli cells and signals through RANK, expressed in most germ cells, whereas the RANKL-inhibitor osteoprotegerin (OPG) is expressed in germ and peritubular cells. OPG treatment increases wild-type mouse sperm counts, and mice with global or Sertoli-specific genetic suppression of Rankl have increased male fertility and sperm counts. Moreover, RANKL levels in seminal fluid are high and distinguishes normal from infertile men with higher specificity than total sperm count. In infertile men, one dose of Denosumab decreases RANKL seminal fluid concentration and increases serum Inhibin-B and anti-Müllerian-hormone levels, but semen quality only in a subgroup. This translational study suggests that RANKL is a regulator of male reproductive function, however, predictive biomarkers for treatment-outcome requires further investigation in placebo-controlled studies.
Peters S, Clézardin P, Márquez-Rodas I, Niepel D, Gedye C. Clin Transl Oncol. 2019;21:977–91. doi: 10.1007/s12094-018-02023-5. Epub 2019 Jan 17. PMID: 30656607.
Abstract
Drug repurposing offers advantages over traditional drug development in terms of cost, speed and improved patient outcomes. The receptor activator of nuclear factor kappa B (RANK) ligand (RANKL) inhibitor denosumab is approved for the prevention of skeletal-related events in patients with advanced malignancies involving bone, including solid tumours and multiple myeloma. Following improved understanding of the role of RANK/RANKL in cancer biology, denosumab has already been repurposed as a treatment for giant cell tumour of bone. Here, we review the role of RANK/RANKL in tumourigenesis, including effects on tumour initiation, progression and metastasis and consider the impact of RANK/RANKL on tumour immunology and immune evasion. Finally, we look briefly at ongoing trials and future opportunities for therapeutic synergy when combining denosumab with anti-cancer agents such as immune checkpoint inhibitors.
Juel Mortensen L, Lorenzen M, Jørgensen N, Andersson A-M, Nielsen JE, Petersen LI, et al.Bone. 2019;123:103–14. doi: 10.1016/j.bone.2019.03.022. Epub 2019 Mar 23. PMID: 30914274.
Power RJ, Hearn J, Gillis CJ, Harvey D, French C, Organ M. Can Urol Assoc J. 2021 Apr;15(4):E221-E226. doi: 10.5489/cuaj.6675. PMID: 33007179; PMCID: PMC8021429.
Globally, about 1 in 6 deaths is related to cancer. Advances in cancer research have improved the prevention, the detection, and the treatment of cancer. Understanding on how cancers starts, grows and spreads is important for cancer treatment. Biomarkers play a critical role at all stages of the disease and serve as therapeutic targets (1).
Biomarkers in Cancer Research
NEUROPILIN-1 is a checkpoint target
The biomarker Neuropilin-1 (NRP1) has gained renewed attention as it is implicated in promoting tumor progression. It acts as a co-receptor to VEGF (Vascular Endothelial Growth Factor) and induces angiogenesis, the process of the formation of new blood-vessels (2). NRP-1 is expressed in a variety of cancers including lung, prostate, pancreas or colon carcinoma. In metastatic melanoma, NRP-1 plays a crucial role in melanoma aggressiveness and evidence supports its use as a target for therapies (3) .
More recently, Neuropilin-1 has been identified as a checkpoint target with unique implications for cancer immunology and immunotherapy (4). This review discusses the increasing literature on Neuropilin-1 mediated immune modulation providing a rationale to categorize NRP1 as a key checkpoint in the tumor microenvironment (TME) as well as a promising immunotherapeutic target.
Did you know: Neuropilin-1 can easily be detected in serum, plasma and in cell culture supernatants using a conventional ELISA kit?
The assay is fully validated for clinical use in human samples but also works in non-human samples. Only 10µl sample is required.
Neuropilin-1: a checkpoint target with unique implications for cancer immunology and immunotherapy
Abstract
Checkpoint blockade immunotherapy established a new paradigm in cancer treatment: for certain patients curative treatment requires immune reinvigoration. Despite this monumental advance, only 20%-30% of patients achieve an objective response to standard of care immunotherapy, necessitating the consideration of alternative targets. Optimal strategies will not only stimulate CD8+ T cells, but concomitantly modulate immunosuppressive cells in the tumor microenvironment (TME), most notably regulatory T cells (Treg cells). In this context, the immunoregulatory receptor Neuropilin-1 (NRP1) is garnering renewed attention as it reinforces intratumoral Treg cell function amidst inflammation in the TME. Loss of NRP1 on Treg cells in mouse models restores antitumor immunity without sacrificing peripheral tolerance. Enrichment of NRP1+ Treg cells is observed in patients across multiple malignancies with cancer, both intratumorally and in peripheral sites. Thus, targeting NRP1 may safely undermine intratumoral Treg cell fitness, permitting enhanced inflammatory responses with existing immunotherapies. Furthermore, NRP1 has been recently found to modulate tumor-specific CD8+ T cell responses. Emerging data suggest that NRP1 restricts CD8+ T cell reinvigoration in response to checkpoint inhibitors, and more importantly, acts as a barrier to the long-term durability of CD8+ T cell-mediated tumor immunosurveillance. These novel and distinct regulatory mechanisms present an exciting therapeutic opportunity. This review will discuss the growing literature on NRP1-mediated immune modulation which provides a strong rationale for categorizing NRP1 as both a key checkpoint in the TME as well as an immunotherapeutic target with promise either alone or in combination with current standard of care therapeutic regimens.
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Citations:
The Biomedica human Angiopoietin-2 ELISA (cat.no. BI-ANG2) is described in Nephrology Dialysis Transplantation, Volume 34, Issue Supplement_1, June 2019, gfz103.SP339:
Ana-Maria Suciu Andreea, Jacqueline Wallwitz, Berg Gabriela, Maria Laber Anna, Himmler Gottfried
INTRODUCTION: Angiopoietin-2 (Ang-2) is an important regulator of the angiopoietin-1/Tie-2 receptor signaling system on endothelial cells during angiogenesis. Disruption of this signaling leads to the loss of endothelial integrity and renders the endothelium response towards a variety of pro-inflammatory cytokines and growth factors. Thus, Ang-2 might lead to vascular micro-inflammation in patients with CKD (chronic kidney disease). Ang-2 levels increase with CKD stage, are associated with fluid overload and abnormal cardiac structure and predict mortality in patients with CKD stages 4–5. Although Ang-2 levels return toward normal after successful kidney transplantation, Ang-2 remains a putative cardiovascular risk factor in this population.
METHODS: An enzyme-linked immunosorbent assay for the detection of all three angiopoietin-2 isoforms in human serum and plasma was developed. Two high quality antibodies are combined in a sandwich test format: As capturing antibody a recombinant monoclonal antibody is used. A biotin-labeled polyclonal affinity-purified antibody serves for detection of the analyte. High resolution epitope mapping of the antibodies via peptide microarray technology allowed the identification of linear antibody epitopes. Technical performance and accuracy of the assay were assessed according to ICH/EMEA guidelines.
RESULTS: Microarray data illustrate the binding of the polyclonal detection antibody to human angiopoietin-2 spotted on a chip. Altogether, seven linear epitopes located N-terminal/at the center of angiopoietin-2 were detected. Most relevant linear epitopes are epitope e2 in the super clustering region and e6 near the fibrinogen-like domain, displaying a twofold higher fluorescent signal than the remaining epitopes. For the recombinant monoclonal antibody no fluorescence was recorded. This antibody recognizes a structural epitope within the C-terminus of angiopoietin-2, which covers the bioactive receptor-binding site of the protein. We expect to detect all described angiopoietin-2 isoforms, as the receptor-binding site is conserved and the majority of the polyclonal antibody epitopes are present in all isoforms. This assay is in conformance with ICH/EMEA/FDA guidelines. Validation data demonstrate its applicability in nephrological disorders including chronic kidney disease. Here we compare an apparently healthy population to chronic kidney disease samples on haemodialysis and kidney transplant samples.
CONCLUSIONS: This new angiopoietin-2 ELISA enables a quick and accurate quantification of all human angiopoietin-2 isoforms that are bioactive in chronic kidney disease and other nephrological disorders.
The Angiopoietin-2 (ANG2) ELISA assay has been fully validated according to international quality guidelines (ICH, EMEA, FDA) and is in line with the required quality specifications (data are shown in the validation file). Assay performance characteristics as accuracy, dilution linearity and parallelism, precision, detection limit and sensitivity, sample stability, specificity including characterization of the angiopoietin-2 antibodies, epitope-mapping were performed.
Osteologie 2020; 29(01): 72. DOI: 10.1055/s-0039-3402888. G Berg, Andreea Ana-Maria Suciu, E Gadermaier, J Wallwitz, G Himmler
Introduction Vascular endothelial growth factor A (VEGF-A), a prominent member of growth factors that regulate angiogenesis and development of normal vasculature, plays an important role in bone development and remodeling. Studies have shown that ossification requires vascularization a priori and that most VEGF in the bone comes from osteoblastic cells. Upon secretion from osteoblasts, VEGF activates endothelial cell migration/proliferation and vessel permeability. Moreover, it regulates osteoclastic differentiation and migration in bone repair. Thus, VEGF represents a relevant therapeutic target. The sensitive measurement of low amounts of circulating VEGF-A found in control cohorts of apparently healthy individuals proves to be difficult. Hence, there is a need for a high-sensitivity assay that reliably measures low VEGF-A concentrations.
Methods We developed a high-sensitivity sandwich ELISA for the detection of human total bioactive VEGF-A using high quality, well-characterized recombinant monoclonal and polyclonal anti-human VEGF-A antibodies. The linear epitopes of the polyclonal detection antibody were mapped with microarray technology. Analyte stability was determined and in accordance with ICH and EMEA guidelines, assay parameters like specificity, dilution linearity, and spike recovery were assessed.
Results We demonstrate that bioactive human VEGF-A can reliably be measured in plasma preparations. In contrast, serum VEGF levels are clearly increased in some samples. This indicates that serum preparation might have an influence on the VEGF amount measured as VEGF can be released from platelets during sample manipulation. Most importantly, we show that samples of apparently healthy individuals are measurable over background. The assay covers a calibration range between 0 and 2000 pg/ml and assay characteristics as well as analyte stability meet the international standards of acceptance. The recombinant capture antibody recognizes a structural epitope in the conserved receptor binding-site of VEGF-A, and thus, specifically binds to all bioactive isoforms of VEGF-A. The polyclonal detection antibody recognizes linear epitopes in the first 120 amino acids of the VEGF-A molecule.
Discussion Our novel VEGF-A ELISA provides a reliable and accurate tool for the quantitative determination of all biologically active VEGF-A isoforms with high sensitivity.
Keywords Angiogenesis, Bone Formation, Growth Factors, Bone Repair, Vascular Endothelial Growth Factor
Of note: The VEGF ELISA assay has been validated according to the guidelines of ICH, EMEA, and FDA. The VEGF assay validation data show that they correspond to the international quality specifications. Various experiments including parallelism and dilution linearity as well as accuracy, precision, analysis of the stability of samples, and the characterization of antibodies utilized in the VEGF ELISA assay e.g. epitope-mapping, were performed.
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Background: Coronary artery ectasia (CAE) is found in about 1% of coronary angiography and is associated with poor clinical outcomes. The prognostic value of plasma big Endothelin-1 (ET-1) in CAE remains unknown. Methods: Patients with angiographically confirmed CAE from 2009 to 2015, who had big ET-1 data available were included. The primary outcome was 5-year major adverse cardiovascular events (MACE), defined as a component of cardiovascular death and non-fatal myocardial infarction (MI). Patients were divided into high or low big ET-1 groups using a cut-off value of 0.58 pmol/L, according to the receiver operating characteristic curve. Kaplan-Meier method, propensity score method, and Cox regression were used to assess the clinical outcomes in the 2 groups. Results: A total of 992 patients were included, with 260 in the high big ET-1 group and 732 in the low big ET-1 group. At 5-year follow-up, 57 MACEs were observed. Kaplan-Meier analysis and univariable Cox regression showed that patients with high big ET-1 levels were at increased risk of MACE (9.87 vs. 4.50%; HR 2.23, 95% CI 1.32-3.78, P = 0.003), cardiovascular death (4.01 vs. 1.69%; HR 2.37, 95% CI 1.02-5.48, P = 0.044), and non-fatal MI (6.09 vs. 2.84%; HR 2.17, 95% CI 1.11-4.24, P = 0.023). A higher risk of MACE in the high big ET-1 group was consistent in the propensity score matched cohort and propensity score weighted analysis. In multivariable analysis, a high plasma big ET-1 level was still an independent predictor of MACE (HR 1.82, 95% CI 1.02-3.25, P = 0.043). A combination of high plasma big ET-1 concentrate and diffuse dilation, when used to predict 5-year MACE risk, yielded a C-statistic of 0.67 (95% CI 0.59-0.74). Conclusion: Among patients with CAE, high plasma big ET-1 level was associated with increased risk of MACE, a finding that could improve risk stratification.