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Rationale & objectiveTubulointerstitial damage is a feature of early chronic kidney disease (CKD), but current clinical tests capture it poorly. Urine biomarkers of tubulointerstitial health may identify risk of CKD.Study designProspective cohort (Atherosclerosis Risk in Communities [ARIC]) and case-cohort (Multi-Ethnic Study of Atherosclerosis [MESA] and Reasons for Geographic and Racial Differences in Stroke [REGARDS]).Setting & participantsAdults with estimated glomerular filtration rate (eGFR) ≥60 mL/min/1.73 m2 and without diabetes in the ARIC, REGARDS, and MESA studies.ExposuresBaseline urine monocyte chemoattractant protein-1 (MCP-1), alpha-1-microglobulin (α1m), kidney injury molecule-1, epidermal growth factor, and chitinase-3-like protein 1.OutcomeIncident CKD or end-stage kidney disease.Analytical approachMultivariable Cox proportional hazards regression for each cohort; meta-analysis of results from all 3 cohorts.Results872 ARIC participants (444 cases of incident CKD), 636 MESA participants (158 cases), and 924 REGARDS participants (488 cases) were sampled. Across cohorts, mean age ranged from 60 ± 10 to 63 ± 8 years, and baseline eGFR ranged from 88 ± 13 to 91 ± 14 mL/min/1.73 m2. In ARIC, higher concentrations of urine MCP-1, α1m, and kidney injury molecule-1 were associated with incident CKD. In MESA, higher concentration of urine MCP-1 and lower concentration of epidermal growth factor were each associated with incident CKD. In REGARDS, none of the biomarkers were associated with incident CKD. In meta-analysis of all 3 cohorts, each 2-fold increase α1m concentration was associated with incident CKD (HR, 1.19; 95% CI, 1.08-1.31).LimitationsObservational design susceptible to confounding; competing risks during long follow-up period; meta-analysis limited to 3 cohorts.ConclusionsIn 3 combined cohorts of adults without prevalent CKD or diabetes, higher urine α1m concentration was independently associated with incident CKD. 4 biomarkers were associated with incident CKD in at least 1 of the cohorts when analyzed individually. Kidney tubule health markers might inform CKD risk independent of eGFR and albuminuria.

Summary: Background: Cardiac surgery invariably triggers acute kidney stress causing adverse renal outcomes. The AKITA study evaluated the efficacy and safety of RMC-035, a novel analogue of alpha-1-microglobulin, for reducing cardiac surgery-associated kidney injury. Methods: In this randomised double-blind placebo-controlled phase 2a study, we randomly assigned (1:1) adult hospitalised patients undergoing open-chest cardiac surgery at high risk for acute kidney injury (AKI) at 21 sites in North America and Europe to receive either RMC-035 (1.3 or 0.65 mg/kg) or placebo (1:1) for 2 days (5 intravenous infusions), stratified by region and renal function. Eligible patients had at least one pre-defined AKI risk factor. Patients with severe renal impairment (estimated glomerular filtration rate [eGFR]

Online hemodiafiltration (OHDF) for renal replacement therapy has two modes: pre- (pre-OHDF) and post-dilution OHDF (post-OHDF). To elucidate the precise differences between the two modes, a clinical study was performed using the same polysulfone hemodiafilters in the same patients. Eight patients were treated with ABH™-22PA for 6 weeks: 3 weeks of pre-OHDF (with substitution volumes of 24, 36, and 48 L) and 3 weeks of post-OHDF (6, 8, and 10 L). The reduction ratios of urea, uric acid (UA), creatinine (CRE), inorganic phosphorus (iP), beta-2-microglobulin (β2-MG), and alpha-1-microglobulin (α1-MG) were evaluated. The removal amounts of β2-MG, α1-MG, and albumin were also evaluated by analyzing the spent dialysis fluids. The types and numbers of adverse events (AEs) and device malfunctions were recorded. The reduction ratios of urea, UA, CRE, iP, and β2-MG were comparable among all conditions, while that of α1-MG tended to be slightly higher in post-OHDF than in pre-OHDF. The removal amounts of α1-MG and albumin in pre-OHDF and post-OHDF were significantly greater with the maximum substitution volume than with the minimum volume. However, the selective removal indices, which were obtained by dividing the amount of α1-MG removed by the albumin level, tended to be slightly higher in pre- than in post-OHDF. No device-related AEs or device malfunctions occurred in either mode. No significant differences in inflammatory responses, evaluated by high-sensitivity C-reactive protein and interleukin-6, were observed. This study provides removal performance and safety data regarding the application of ABH-22PA for pre- and post-OHDF. [ABSTRACT FROM AUTHOR]

Introduction: Acute kidney injury (AKI) is a common complication in cardiac surgery patients and prevention is needed to improve clinical outcomes. Alpha-1-microglobulin (A1M) is a physiological antioxidant with strong tissue-protective and cell-protective properties that has demonstrated renoprotective effects. RMC-035, a recombinant variant of endogenous human A1M, is being developed for the prevention of AKI in cardiac surgery patients. Methods: In this phase 1b, randomized, double-blind, and parallel group clinical study, 12 cardiac surgery patients undergoing elective, open-chest, on-pump coronary artery bypass graft and/or valve surgery with additional predisposing AKI risk factors were enrolled to receive in total 5 intravenous doses of either RMC-035 or placebo. The primary objective was to evaluate the safety and tolerability of RMC-035. The secondary objective was to evaluate its pharmacokinetic properties. Results: RMC-035 was well tolerated. The nature and frequency of adverse events (AEs) were consistent with the expected background rates in the underlying patient population with no AEs reported as related to study drug. No clinically relevant changes were observed for vital signs or laboratory parameters except for renal biomarkers. Several established AKI urine biomarkers were reduced at 4 hours after first dose administration in the treatment group, indicating a reduced perioperative tubular cell injury following RMC-035 treatment. Conclusion: Multiple intravenous doses of RMC-035 were well tolerated in patients undergoing cardiac surgery. Observed RMC-035 plasma exposures were safe and in the range of expected pharmacological activity. Furthermore, urine biomarkers suggest reduced perioperative kidney cell injury, warranting further investigation of RMC-035 as a potential renoprotective treatment.

Cardiac hepatopathy refers to acute or chronic liver damage caused by cardiac dysfunction in the absence of any other possible causative reasons of liver injury. There is a large number of evidence of the fact that cardiac hepatopathy is associated with poor clinical outcomes in patients with acute or actually decompensated heart failure (HF). However, the currently dominated pathophysiological background does not explain a role of metabolic regulative proteins secreted by hepatocytes in progression of HF, including adverse cardiac remodeling, kidney injury, skeletal muscle dysfunction, osteopenia, sarcopenia and cardiac cachexia. The aim of this narrative review was to accumulate knowledge of hepatokines (adropin; fetuin-A, selenoprotein P, fibroblast growth factor-21, and alpha-1-microglobulin) as adaptive regulators of metabolic homeostasis in patients with HF. It is suggested that hepatokines play a crucial, causative role in inter-organ interactions and mediate tissue protective effects counteracting oxidative stress, inflammation, mitochondrial dysfunction, apoptosis and necrosis. The discriminative potencies of hepatokines for HF and damage of target organs in patients with known HF is under on-going scientific discussion and requires more investigations in the future. [ABSTRACT FROM AUTHOR]

BACKGROUND:HIV-infected (HIV+) persons are at increased risk of chronic kidney disease, but serum creatinine does not detect early losses in kidney function. We hypothesized that urine biomarkers of kidney damage would be associated with subsequent changes in kidney function in a contemporary cohort of HIV+ and HIV-uninfected (HIV-) men. METHODS:In the Multicenter AIDS Cohort Study, we measured baseline urine concentrations of 5 biomarkers from 2009 to 2011 in 860 HIV+ and 337 HIV- men: albumin, alpha-1-microglobulin (α1m), interleukin-18 (IL-18), kidney injury molecule-1 (KIM-1), and procollagen type III N-terminal propeptide (PIIINP). We evaluated associations of urine biomarker concentrations with annual changes in estimated glomerular filtration rate (eGFR) using multivariable linear mixed models adjusted for demographics, traditional kidney disease risk factors, HIV-related risk factors, and baseline eGFR. RESULTS:Over a median follow-up of 4.8 years, the average annual eGFR decline was 1.42 mL/min/1.73 m2/year in HIV+ men and 1.22 mL/min/1.73 m2/year in HIV- men. Among HIV+ men, the highest vs. lowest tertiles of albumin (-1.78 mL/min/1.73 m2/year, 95% CI -3.47 to -0.09) and α1m (-2.43 mL/min/1.73 m2/year, 95% CI -4.14 to -0.73) were each associated with faster annual eGFR declines after multivariable adjustment. Among HIV- men, the highest vs. lowest tertile of α1m (-2.49 mL/min/1.73 m2/year, 95% CI -4.48 to -0.50) was independently associated with faster annual eGFR decline. Urine IL-18, KIM-1, and PIIINP showed no independent associations with eGFR decline, regardless of HIV serostatus. CONCLUSIONS:Among HIV+ men, higher urine albumin and α1m are associated with subsequent declines in kidney function, independent of eGFR.

Background: Approximately 180 mothers are treated in Swedish Intensive Care Units (ICU) due to preeclampsia each year. Although several clinical risk factors are known, prediction of critical disease is challenging. Two scavenger proteins, hemopexin (Hpx) and alpha-1-microglobulin (A1M) have been suggested to be associated with the oxidative stress seen in preeclampsia. The ratio of two other biomarkers, soluble fms-like tyrosine kinase (sFlt-1) to placental growth factor (PIGF), is predictive of adverse pregnancy outcomes.Methods: In total 121 women were included in this study where we compared risk factors for preeclampsia, plasma levels of Hpx and A1M in ICU-patients with preeclampsia (n = 41) compared to uncomplicated preeclampsia cases (n = 40) and normotensive pregnancies (n = 40), with the objective to identify clinical risk patterns for severe disease. The sFlt-1/PIGF ratio was investigated in early and late onset preeclampsia ICU-patients. Blood samples were collected at admission to ICU and within 27 h postpartum for all groups.Results: Hemopexin and A1M levels were significantly lower in the preeclampsia ICU-cohort compared to uncomplicated preeclampsia patients. The sFlt-1/PIGF-ratio was elevated in the ICU-patients but there was no difference between early and late onset preeclampsia. The ICU-patients had more clinical risk factors, refractory hypertension, and an increased rate of emergency Caesarean section.Conclusion: Intensive care patients have more clinical risk factors and a Hpx and A1M profile suggestive of depletion and thereby a reduced capacity to respond to oxidative stress. The ratios of sFlt-1/PIGF were high in the ICU-cohort and in accordance with pre-delivery levels predictive of adverse pregnancy outcomes. [ABSTRACT FROM AUTHOR]

Introduction: Urine alpha-1-microglobulin (Uα1m) elevations signal proximal tubule dysfunction. In ambulatory settings, higher Uα1m is associated with acute kidney injury (AKI), progressive chronic kidney disease (CKD), cardiovascular (CV) events, and mortality. We investigated the associations of pre- and postoperative Uα1m concentrations with adverse outcomes after cardiac surgery. Methods: In 1,464 adults undergoing cardiac surgery in the prospective multicenter Translational Research Investigating Biomarker Endpoints for Acute Kidney Injury (TRIBE-AKI) cohort, we measured the pre-and postoperative Uα1m concentrations and calculated the changes from pre- to postoperative concentrations. Outcomes were postoperative AKI during index hospitalization and longitudinal risks for CKD incidence and progression, CV events, and all-cause mortality after discharge. We analyzed Uα1m continuously and categorically by tertiles using multivariable logistic regression and Cox proportional hazards regression adjusted for demographics, surgery characteristics, comorbidities, baseline estimated glomerular filtration rate, urine albumin, and urine creatinine. Results: There were 230 AKI events during cardiac surgery hospitalization; during median 6.7 years of follow-up, there were 212 cases of incident CKD, 54 cases of CKD progression, 269 CV events, and 459 deaths. Each 2-fold higher concentration of preoperative Uα1m was independently associated with AKI (adjusted odds ratio [aOR] = 1.36, 95% confidence interval 1.14–1.62), CKD progression (adjusted hazard ratio [aHR] = 1.46, 1.04–2.05), and all-cause mortality (aHR = 1.19, 1.06–1.33) but not with incident CKD (aHR = 1.21, 0.96–1.51) or CV events (aHR = 1.01, 0.86–1.19). Postoperative Uα1m was not associated with AKI (aOR per 2-fold higher = 1.07, 0.93–1.22), CKD incidence (aHR = 0.90, 0.79–1.03) or progression (aHR = 0.79, 0.56–1.11), CV events (aHR = 1.06, 0.94–1.19), and mortality (aHR = 1.01, 0.92–1.11). Conclusion: Preoperative Uα1m concentrations may identify patients at high risk of AKI and other adverse events after cardiac surgery, but postoperative Uα1m concentrations do not appear to be informative. [ABSTRACT FROM AUTHOR]

Abstract Background Preeclampsia remains a significant danger to both mother and child and current prevention and treatment management strategies are limited. The objective of this systematic review was to investigate the current literature on evidence for the use of the regenerative capacity of mesenchymal stem cell (MSC) therapy, the anticoagulant activity of antithrombin (AT), or the free radical scavenging activity of alpha-1-microglobulin (A1M) as potential novel treatments for severe preeclampsia and Hemolysis, Elevated Liver enzymes, Low Platelet count (HELLP). Method We conducted a systematic review of potential biological therapies for preeclampsia. We screened MEDLINE and Embase from inception through May 2017 for studies using AT, A1M or MSCs as potential treatments for preeclampsia and/or HELLP. A meta-analysis was performed to pool data from randomized control trials (RCTs) with homogenous outcomes using the inverse variance method. The Newcastle-Ottawa Scale, the Cochrane risk of bias tool for RCTs, and SYRCLE’s risk of bias tool for animal studies were used to investigate potential bias of studies. Results The literature search retrieved a total of 1015 articles, however, only 17 studies met the selection criteria: AT (n = 9, 8 human and 1 animal); A1M (n = 4, 3 animal and 1 ex-vivo); and, MSCs (n = 4, 3 animal and 1 ex-vivo). A meta-analysis of AT therapy versus placebo and a meta-analysis for AT therapy with heparin versus heparin alone did not show significant differences between study groups. Animal and ex-vivo studies demonstrated significant benefits in relevant outcomes for A1M and MSCs versus control treatments. Most RCT studies were rated as having a low risk of bias across categories with some studies showing an unclear risk of bias in some categories. The two cohort studies both received a total of four out of nine stars (a rating of “poor” quality). Most animal studies had an unclear risk of bias across most categories, with some studies having a low risk of bias in some categories. Conclusions The findings of this review are strengthened by rigorous systematic search and review of the literature. Results of our meta-analyses do not currently warrant further exploration of AT as a treatment of preeclampsia in human trials. Results of animal and ex-vivo studies of A1M and MSCs were encouraging and supportive of initiating human investigations.

Accumulation of damaged or misfolded proteins resulted from oxidative protein modification induces endoplasmic reticulum (ER) stress by activating the pathways of unfolded protein response. In pathologic hemolytic conditions, extracellular free hemoglobin is submitted to rapid oxidation causing heme release. Resident cells of atherosclerotic lesions, after intraplaque hemorrhage, are exposed to heme leading to oxidative injury. Therefore, we raised the question whether heme can also provoke ER stress. Smooth muscle cells are one of the key players of atherogenesis; thus, human aortic smooth muscle cells (HAoSMCs) were selected as a model cell to reveal the possible link between heme and ER stress. Using immunoblotting, quantitative polymerase chain reaction and immunocytochemistry, we quantitated the markers of ER stress. These were: phosphorylated eIF2α, Activating transcription factor-4 (ATF4), DNA-damage-inducible transcript 3 (also known as C/EBP homology protein, termed CHOP), X-box binding protein-1 (XBP1), Activating transcription factor-6 (ATF6), GRP78 (glucose-regulated protein, 78kDa) and heme responsive genes heme oxygenase-1 and ferritin. In addition, immunohistochemistry was performed on human carotid artery specimens from patients who had undergone carotid endarterectomy. We demonstrate that heme increases the phosphorylation of eiF2α in HAoSMCs and the expression of ATF4. Heme also enhances the splicing of XBP1 and the proteolytic cleavage of ATF6. Consequently, there is up-regulation of target genes increasing both mRNA and protein levels of CHOP and GRP78. However, TGFβ and collagen type I decreased. When the heme binding proteins, alpha-1-microglobulin (A1M) and hemopexin (Hpx) are present in cell media, the ER stress provoked by heme is inhibited. ER stress pathways are also retarded by the antioxidant N-acetyl cysteine (NAC) indicating that reactive oxygen species are involved in heme-induced ER stress. Consistent with these findings, elevated expression of the ER stress marker GRP78 and CHOP were observed in smooth muscle cells of complicated lesions with hemorrhage compared to either atheromas or healthy arteries. In conclusion, heme triggers ER stress in a time- and dose-dependent manner in HAoSMCs. A1M and Hpx as well as NAC effectively hamper heme-induced ER stress, supporting their use as a potential therapeutic approach to reverse such a deleterious effects of heme toxicity.

Aims: Human α1-microglobulin (A1M) is an endogenous reductase and radical- and heme-binding protein with physiological antioxidant protective functions. Recombinant human A1M (rA1M) has been shown to have therapeutic properties in animal models of preeclampsia, a pregnancy disease associated with oxidative stress. Recombinant A1M, however, lacks glycosylation, and shows lower solubility and stability than A1M purified from human plasma. The aims of this work were to (i) use site-directed mutagenesis to improve the physicochemical properties of rA1M, (ii) demonstrate that the physicochemically improved rA1M displays full in vitro cell protective effects as recombinant wild-type A1M (rA1M-wt), and (iii) show its therapeutic potential in vivo against acute kidney injury (AKI), another disease associated with oxidative stress. Results: A novel recombinant A1M-variant (rA1M-035) with three amino acid substitutions was constructed, successfully expressed, and purified. rA1M-035 had improved solubility and stability compared with rA1M-wt, and showed intact in vitro heme-binding, reductase, antioxidation, and cell protective activities. Both rA1M-035 and rA1M-wt showed, for the first time, potential in vivo protective effects on kidneys using a mouse rhabdomyolysis glycerol injection model of AKI. Innovation: A novel recombinant A1M-variant, rA1M-035, was engineered. This protein showed improved solubility and stability compared with rA1M-wt, full in vitro functional activity, and potential protection against AKI in an in vivo rhabdomyolysis mouse model. Conclusion: The new rA1M-035 is a better drug candidate than rA1M-wt for treatment of AKI and preeclampsia in human patients. [ABSTRACT FROM AUTHOR]

Accumulation of damaged or misfolded proteins resulted from oxidative protein modification induces endoplasmic reticulum (ER) stress by activating the pathways of unfolded protein response. In pathologic hemolytic conditions, extracellular free hemoglobin is submitted to rapid oxidation causing heme release. Resident cells of atherosclerotic lesions, after intraplaque hemorrhage, are exposed to heme leading to oxidative injury. Therefore, we raised the question whether heme can also provoke ER stress. Smooth muscle cells are one of the key players of atherogenesis; thus, human aortic smooth muscle cells (HAoSMCs) were selected as a model cell to reveal the possible link between heme and ER stress. Using immunoblotting, quantitative polymerase chain reaction and immunocytochemistry, we quantitated the markers of ER stress. These were: phosphorylated eIF2α, Activating transcription factor-4 (ATF4), DNA-damage-inducible transcript 3 (also known as C/EBP homology protein, termed CHOP), X-box binding protein-1 (XBP1), Activating transcription factor-6 (ATF6), GRP78 (glucose-regulated protein, 78kDa) and heme responsive genes heme oxygenase-1 and ferritin. In addition, immunohistochemistry was performed on human carotid artery specimens from patients who had undergone carotid endarterectomy. We demonstrate that heme increases the phosphorylation of eiF2α in HAoSMCs and the expression of ATF4. Heme also enhances the splicing of XBP1 and the proteolytic cleavage of ATF6. Consequently, there is up-regulation of target genes increasing both mRNA and protein levels of CHOP and GRP78. However, TGFβ and collagen type I decreased. When the heme binding proteins, alpha-1-microglobulin (A1M) and hemopexin (Hpx) are present in cell media, the ER stress provoked by heme is inhibited. ER stress pathways are also retarded by the antioxidant N-acetyl cysteine (NAC) indicating that reactive oxygen species are involved in heme-induced ER stress. Consistent with these findings, elevated expression of the ER stress marker GRP78 and CHOP were observed in smooth muscle cells of complicated lesions with hemorrhage compared to either atheromas or healthy arteries. In conclusion, heme triggers ER stress in a time- and dose-dependent manner in HAoSMCs. A1M and Hpx as well as NAC effectively hamper heme-induced ER stress, supporting their use as a potential therapeutic approach to reverse such a deleterious effects of heme toxicity. [ABSTRACT FROM AUTHOR]

Introduction:Pyelonephritis is known as kidney inflammation due to bacterial infection which should be diagnosed and treated promptly. In this article, we decided to systematically review the diagnostic value and reliability of evaluating urine excretion low molecular weight protein alpha-1-microglobulin (A1M). Methods: PubMed and Scopus were searched for the relevant articles about the efficacy of urine alpha-1-micriglobulin assays in the diagnosis of pyelonephritis in children. The search strategy was microglobulin AND pyelonephritis. No language and date limitations were included in this review. Results: A total of 16 articles were retrieved from PubMed and 23 articles from Scopus. After studying the abstracts, only 5 articles were selected, which specifically studied the efficacy of alpha-1-micrglobulin in the diagnosis of pyelonephritis in children.Discussion: A1M is not an acute phase protein but its concentration alters in several clinical conditions. Conclusion: Evaluating the urine concentration of A1M is a noninvasive and cost effective strategy with the diagnostic capability for urinary tract disorders such as early recognition of tubular damages during pyelonephritis.

Background Urine cadmium (Cd) and renal function biomarkers, mostly analysed in urine spot samples, are well established biomarkers of occupational exposure. Their use and associations at low environmental level are common, but have recently been questioned, particularly in terms of physiological variability and normalisation bias in the case of urine spot samples. Aim To determine the appropriateness of spot urine and/or blood Cd exposure biomarkers and their relationships with renal function biomarkers at low levels of exposure. To this end, we used data from Slovenian human biomonitoring program involving 1081 Slovenians (548 males, mean age 31 years; 533 lactating females, mean age 29 years; 2007–2015) who have not been exposed to Cd occupationally. Results Geometric means (GMs) of Cd in blood and spot urine samples were 0.27 ng/mL (0.28 for males and 0.33 for females) and 0.19 ng/mL (0.21 for males and 0.17 for females), respectively. Differing results were obtained when contrasting normalisation by urine creatinine with specific gravity. GMs of urine albumin (Alb), alpha-1-microglobulin (A1M), N-acetyl-beta-glucosaminidase (NAG), and immunoglobulin G (IgG) were far below their upper reference limits. Statistical analysis of unnormalised or normalised urine data often yielded inconsistent and conflicting results (or trends), so association analyses with unnormalised data were taken as more valid. Relatively weak positive associations were observed between urine Cd (ng/mL) and blood Cd (β=0.11, p=0.002 for males and β=0.33, p<0.001 for females) and for females between urine NAG and blood Cd (β=0.14, p=0.04). No associations were found between other renal function biomarkers and blood Cd. Associations between Cd and renal function biomarkers in urine were stronger (p<0.05, β=0.11–0.63). Mostly, all of the associations stayed significant but weakened after normalisation for diuresis. In the case of A1M, its associations with Cd were influenced by current smoking and blood Pb in males and by pre-pregnancy smoking and blood Se in females (β up to 0.34, p<0.001). Statistical analysis of unnormalised or normalised urine data often yielded inconsistent and conflicting results (or trends), so association analyses data with unnormalised were taken as more valid. Conclusions The observed uncertainties introduced by urine normalisation, particularly by creatinine, confirm blood Cd as a superior low-Cd exposure biomarker versus urine Cd in cases when 24 h urine is unattainable. Evidence that A1M can be positively related to Cd, smoking (current or pre-pregnancy), Pb, and Se status, points to the versatile biological functions of A1M. [ABSTRACT FROM AUTHOR]

Introduction: Acute kidney injury (AKI) is a common complication in cardiac surgery patients and prevention is needed to improve clinical outcomes. Alpha-1-microglobulin (A1M) is a physiological antioxidant with strong tissue-protective and cell-protective properties that has demonstrated renoprotective effects. RMC-035, a recombinant variant of endogenous human A1M, is being developed for the prevention of AKI in cardiac surgery patients., Methods: In this phase 1b, randomized, double-blind, and parallel group clinical study, 12 cardiac surgery patients undergoing elective, open-chest, on-pump coronary artery bypass graft and/or valve surgery with additional predisposing AKI risk factors were enrolled to receive in total 5 intravenous doses of either RMC-035 or placebo. The primary objective was to evaluate the safety and tolerability of RMC-035. The secondary objective was to evaluate its pharmacokinetic properties., Results: RMC-035 was well tolerated. The nature and frequency of adverse events (AEs) were consistent with the expected background rates in the underlying patient population with no AEs reported as related to study drug. No clinically relevant changes were observed for vital signs or laboratory parameters except for renal biomarkers. Several established AKI urine biomarkers were reduced at 4 hours after first dose administration in the treatment group, indicating a reduced perioperative tubular cell injury following RMC-035 treatment., Conclusion: Multiple intravenous doses of RMC-035 were well tolerated in patients undergoing cardiac surgery. Observed RMC-035 plasma exposures were safe and in the range of expected pharmacological activity. Furthermore, urine biomarkers suggest reduced perioperative kidney cell injury, warranting further investigation of RMC-035 as a potential renoprotective treatment., (© 2023 Published by Elsevier Inc. on behalf of the International Society of Nephrology.)

It is generally accepted that development of novel therapies to treat pregnancy-relates disorders, such as preeclampsia, is hampered to the paucity of research funding. Hence, it is with great interest to become aware of at least three novel therapeutic approaches for the treatment of this disorder, exploiting either the anticoagulant activity of antithrombin, the free radical scavenging activity of alpha-1-microglobulin, or the regenerative capacity of placenta-derived mesenchymal stem cells. As these projects are being carried out by small biotech enterprises, the question arises of how they are able to fund such undertakings. A novel strategy adopted by two of these companies is that they successfully petitioned US and EU agencies in order that preeclampsia be accepted in the register of rare or orphan diseases. This provides a number of benefits including market exclusivity, assistance with clinical trials and dedicated funding schemes. Other strategies to supplement meager research funds, especially to test novel approaches, could be crowdfunding, a venture which relies on intimate interaction with advocacy groups. In other words, preeclampsia meets Facebook. Perhaps similar strategies can be adopted to examine novel therapies targeting either the imbalance in angiogenic growth factors, complement activation, reduced levels of placenta protein 13 or excessive neutrophil activation evident in preeclampsia.

Preeclampsia is a leading cause of pregnancy complications and affects 3–7 % of pregnant women. This review summarizes the current knowledge of a new potential etiology of the disease, with a special focus on hemoglobin-induced oxidative stress. Furthermore, we also suggest hemoglobin as a potential target for therapy. Gene and protein profiling studies have shown increased expression and accumulation of free fetal hemoglobin in the preeclamptic placenta. Predominantly due to oxidative damage to the placental barrier, fetal hemoglobin leaks over to the maternal circulation. Free hemoglobin and its metabolites are toxic in several ways; a) ferrous hemoglobin (Fe2+) binds strongly to the vasodilator nitric oxide and reduces the availability of free nitric oxide, which results in vasoconstriction, b) hemoglobin (Fe2+) with bound oxygen spontaneously generates free oxygen radicals and c) the heme groups create an inflammatory response by inducing activation of neutrophils and cytokine production. The endogenous protein α1-microglobulin, with radical and heme binding properties, has shown both ex vivo and in vivo to have the ability to counteract free hemoglobin-induced placental and kidney damage. Oxidative stress in general, and more specifically fetal hemoglobin-induced oxidative stress, could play a key role in the pathology of preeclampsia seen both in the placenta and ultimately in the maternal endothelium.

Background: Alpha-1-microglobulin (A1M), a small lipocalin protein found in plasma and tissues, has been identified as a heme and radical scavenger that may participate in the mitigation of toxicities caused by degradation of hemoglobin. The objective of this work was to investigate heme interactions with A1M in vitro using various analytical techniques and to optimize analytical methodology suitable for rapid evaluation of the ligand binding properties of recombinant A1M versions. Methods: To examine heme binding properties of A1M we utilized UV/Vis absorption spectroscopy, visible circular dichroism (CD), catalase-like activity, migration shift electrophoresis, and surface plasmon resonance (SPR), which was specifically developed for the assessment of His-tagged A1M. Results: The results of this study confirm that A1M is a heme binding protein that can accommodate heme at more than one binding site and/or in coordination with different amino acid residues depending upon heme concentration and ligand-to-protein molar ratio. UV/Vis titration of A1M with heme revealed an unusually large bathochromic shift, up to 38 nm, observed for heme binding to a primary binding site. UV/Vis spectroscopy, visible CD and catalase-like activity suggested that heme is accommodated inside His-tagged (tgA1M) and tagless A1M (ntA1M) in a rather similar fashion although the His-tag is very likely involved into coordination with iron of the heme molecule. SPR data indicated kinetic rate constants and equilibrium binding constants with KD values in a uM range. Conclusions: This study provided experimental evidence of the A1M heme binding properties by aid of different techniques and suggested an analytical methodology for a rapid evaluation of ligand-binding properties of recombinant A1M versions, also suitable for other His-tagged proteins.

Background: Medication-overuse headache (MOH) is a chronic disorder that results from the overuse of analgesics drugs, triptans or other acute headache compounds. Although the exact mechanisms underlying MOH remain still unknown, several studies suggest that it may be associated with development of 'central sensitization', which may cause cutaneous allodynia (CA). Furthermore, the epidemiology of drug-induced disorders suggests that medication overuse could lead to nephrotoxicity. The aim of this work was to confirm and validate the results obtained from previous proteomics studies, in which we analyzed the urinary proteome of MOH patients in comparison with healthy non-abusers individuals. Methods: MOH patients were divided into groups on the basis of the drug abused: triptans, non-steroidal anti-inflammatory drugs (NSAIDs) and mixtures, (mainly containing indomethacin, paracetamol and, in some cases, caffeine). Healthy subjects, with a history of normal renal function, were used as controls. In this study, four proteins that were found differentially expressed in urine, and, on the basis of the literature review, resulted related to kidney diseases, were verified by Western Blot and Enzyme-linked Immunosorbent Assay (ELISA); Prostaglandin-H2 D-synthase (PTGDS), uromodulin (UROM), alpha-1-microglobulin (AMBP) and cystatin-C (CYSC). Results: Western blot analysis allowed to validate our previous proteomics data, confirming that all MOH patients groups show a significant over-excretion of urinary PTGDS, UROM, AMBP and CYSC (excluding triptans group for this latter), in comparison with controls. Moreover, the expression of PTGDS was further evaluated by ELISA. Also by this assay, a significant increase of PTGDS was observed in all MOH abusers, according to 2-DE and Western blot results. Conclusions: In this study, we confirmed previous findings concerning urinary proteins alterations in MOH patients, identified and demonstrated the over-expression of PTGDS, UROM, AMBP, and CYSC, particularly in NSAIDs and mixtures abusers. Over-expression of these proteins have been related to renal dysfunction and probably, PTGDS, to the development of CA. The detection and confirmation of this proteins pattern represent a promising tool for a better understanding of potential nephrotoxicity induced by drugs overuse and may enhance awareness related to the MOH-associated risks, even in absence of clinical symptoms. [ABSTRACT FROM AUTHOR]

Introduction and aims: Balkan endemic nephropathy (BEN), a regional tubulointerstitial kidney disease encountered in South-Eastern Europe, with still undefined etiology and inexorable evolution towards end stage renal disease, raises the question of the relative contribution of family and environmental factors in its etiology. In order to evaluate the intervention of these factors, markers of tubular injury have been assessed, this lesion being considered an early renal involvement in BEN. Methods: The paper studies relatives of BEN patients currently included in dialysis programmes (for involvement of the family factor) and their neighbors (for involvement of environmental factors) and analyzes them with regard to tubular injury by means of tubular biomarkers ( N-acetyl-beta- d-glucosaminidase-NAG and alpha-1-microglobulin), and albuminuria. At the same time, glomerular filtration rate (GFR) (CKD-EPI) was measured. It is considered that, in order to acquire the disease, one should have lived for 20 years in the BEN area. The relatives have been classified according to this criterion. Results: More evident tubular injury was found in the neighbors of BEN patients living for more than 20 years in the endemic area, which argues in favor of environmental factors. Higher levels of urinary alpha-1-microglobulin and albumin in relatives of BEN patients who had been living for more than 20 years in the area than in relatives with a residence under 20 years, plead for the same hypothesis. GFR was lower in persons who had been living for more than 20 years in the BEN area (neighbors and relatives). Conclusions: Environmental factors could be more important in BEN than family factors. [ABSTRACT FROM AUTHOR]

Preeclampsia is a leading cause of pregnancy complications and affects 3-7% of pregnant women. This review summarizes the current knowledge of a new potential etiology of the disease, with a special focus on hemoglobin-induced oxidative stress. Furthermore, we also suggest hemoglobin as a potential target for therapy. Gene and protein profiling studies have shown increased expression and accumulation of free fetal hemoglobin in the preeclamptic placenta. Predominantly due to oxidative damage to the placental barrier, fetal hemoglobin leaks over to the maternal circulation. Free hemoglobin and its metabolites are toxic in several ways; (a) ferrous hemoglobin (Fe2+) binds strongly to the vasodilator nitric oxide (NO) and reduces the availability of free NO, which results in vasoconstriction, (b) hemoglobin (Fe2+) with bound oxygen spontaneously generates free oxygen radicals, and (c) the heme groups create an inflammatory response by inducing activation of neutrophils and cytokine production. The endogenous protein α1-microglobulin, with radical and heme binding properties, has shown both ex vivo and in vivo to have the ability to counteract free hemoglobin-induced placental and kidney damage. Oxidative stress in general, and more specifically fetal hemoglobin-induced oxidative stress, could play a key role in the pathology of preeclampsia seen both in the placenta and ultimately in the maternal endothelium. [ABSTRACT FROM AUTHOR]

Purpose: Early diagnosis of acute rejection and effective immunosuppressive therapy lead to improvement in graft survival following kidney transplantation. In this study, we aimed to establish a urinary protein profile suitable to distinguish between patients with rejection and stable graft function and to predict acute rejection based on postoperatively collected urine samples. A further objective was to identify candidate proteins for the use as biomarkers in clinical practice. Methods: Urine samples of 116 kidney recipients were included. Rejection was proven by biopsy ( n = 58), and stable transplant function was monitored for at least 2 years ( n = 58). Postoperative urine samples were collected between 3rd and 10th day following transplantation. Urinary protein profiles were obtained by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry. Protein identification and validation were performed using multiplex fluorescence 2DE, peptide mass fingerprinting and enzyme-linked immunosorbent assay. Results: A protein profile including four mass peaks differentiated acute rejection from stable transplants at the time point of rejection and at the postoperative state with 73 % sensitivity and 88 % specificity. Alpha-1-microglobulin (A1MG) and Haptoglobin (Hp) were identified as putative rejection biomarkers. Protein levels were significantly higher in postoperative urine from patients with rejection (A1MG 29.13 vs. 22.06 μg/ml, p = 0.001; Hp 628.34 vs. 248.57 ng/ml, p = 0.003). The combination of both proteins enabled the diagnosis of early rejection with 85 % sensitivity and 80 % specificity. Conclusion: Protein profiling using mass spectrometry is suitable for noninvasive detection of rejection-specific changes following kidney transplantation. A specific protein profile enables the prediction of early acute allograft rejection in the immediate postoperative period. A1MG and Hp appear to be reliable rejection biomarkers. [ABSTRACT FROM AUTHOR]

Background: Alpha-1-microglobulin (A1M), a small lipocalin protein found in plasma and tissues, has been identified as a heme1 and radical scavenger that may participate in the mitigation of toxicities caused by degradation of hemoglobin. The objective of this work was to investigate heme interactions with A1M in vitro using various analytical techniques and to optimize analytical methodology suitable for rapid evaluation of the ligand binding properties of recombinant A1M versions. Methods: To examine heme binding properties of A1M we utilized UV/Vis absorption spectroscopy, visible circular dichroism (CD), catalase-like activity, migration shift electrophoresis, and surface plasmon resonance (SPR), which was specifically developed for the assessment of His-tagged A1M. Results: The results of this study confirm that A1M is a heme binding protein that can accommodate heme at more than one binding site and/or in coordination with different amino acid residues depending upon heme concentration and ligand-to-protein molar ratio. UV/Vis titration of A1M with heme revealed an unusually large bathochromic shift, up to 38 nm, observed for heme binding to a primary binding site. UV/Vis spectroscopy, visible CD and catalase-like activity suggested that heme is accommodated inside His-tagged (tgA1M) and tagless A1M (ntA1M) in a rather similar fashion although the His-tag is very likely involved into coordination with iron of the heme molecule. SPR data indicated kinetic rate constants and equilibrium binding constants with KD values in a μM range. Conclusions: This study provided experimental evidence of the A1M heme binding properties by aid of different techniques and suggested an analytical methodology for a rapid evaluation of ligand-binding properties of recombinant A1M versions, also suitable for other His-tagged proteins. [ABSTRACT FROM AUTHOR]

Purpose: To explore oxidative stress and the radical scavenger α-microglobulin (A1M) in the vitreous body of human eyes with primary rhegmatogenous retinal detachment (RRD). Methods: Levels of carbonyl groups, a marker of oxidative stress, and A1M were measured by ELISA and RIA in 14 vitreous samples derived from patients suffering from RRD, and compared with 14 samples from macula hole (MH) patients. Carbonyl group and A1M levels in RRD samples were statistically related to detachment characteristics. Analysis of total protein level, SDS-PAGE, and Western blotting of A1M was also performed. In a separate experiment, mRNA expression of A1M was measured by RT-PCR in rat retina explants. Results: Levels of carbonyl groups and A1M varied widely in RRD vitreous samples, but were significantly higher in samples derived from eyes with large detachment area and macula-off status, while the presence of vitreous hemorrhage did not show any significant correlation. Compared with MH samples, RRD samples displayed significantly higher levels of A1M, whereas changes in total protein levels and carbonyl groups were not significant. Novel forms of A1M, not previously seen in plasma, were found in the vitreous body by Western blotting. Furthermore, A1M expression was seen in rat retina explants and was upregulated after 24 h of culturing. Conclusion: Oxidative stress is a prominent feature of human eyes with primary RRD, and is directly related to detachment severity. Affected eyes can launch a protective response in the form of the radical scavenger A1M possibly derived from the retina. The results thus indicate potential therapeutic cell loss prevention in RRD by employing the endogeneous radical scavenger A1M. [ABSTRACT FROM AUTHOR]

Background: Associations between cadmium (Cd) and kidney function have been reported even at low levels of exposure in the general population. Recently, the causality of these associations has been questioned. Objectives: We examined associations between urinary Cd (U-Cd; a biomarker of exposure) and urinary proteins that are used as biomarkers of kidney effects, based on repeated short-term sampling in healthy subjects. Methods: Twenty-four hour urine samples were collected on 2 separate days at six fixed times from 30 healthy nonsmoking men and women (median age 39 years). We analyzed the samples (N = 354) for Cd (i.e., U-Cd) and two proteins used as kidney function biomarkers: urinary albumin (U-Alb) and alpha-1-microglobulin (U-A1M). Concentrations were adjusted for creatinine concentration or for specific gravity, and excretion rates (mass per hour) were calculated. Possible associations were assessed within each individual participant, and mean correlations and regressions were evaluated. Results: We found clear positive mean associations within individuals between the excretion of U-Cd [mean, 0.11 μg/g creatinine (range, 0.01–0.52 μg/g creatinine)] and both U-Alb and U-A1M. The associations were stronger for excretion rates and concentrations adjusted for specific gravity than for concentrations adjusted for creatinine. We also found significant positive associations of urinary flow with excretion of U-Cd, U-Alb, and U-A1M. Conclusions: Associations between short-term changes in U-Cd and markers of kidney function within individual nonsmoking study participants are unlikely to reflect effects of Cd toxicity. A more likely explanation is that these associations result from normal variation in renal function, including changes in urinary flow, that influence the urinary excretion of both Cd and proteins in the same direction. These effects of normal variability may result in overestimation of the adverse effects of Cd on kidney function at low-level Cd exposure. [ABSTRACT FROM AUTHOR]

Proteinuria is a common complication after renal transplantation (RTx). In adults, tubular proteinuria prevails and is associated with impaired graft survival. In the absence of studies on proteinuria profiling in transplanted children, we aimed at analyzing the types of proteinuria in transplanted children. Fifty-three children (11.8 years) were analyzed in a cross-sectional study. Morning urine was tested for total protein (PROT), albumin (ALB) and alpha-1-microglobulin (AMG). The type of proteinuria was assessed by the alpha-1-microglobulin/albumin algorithm (AAA): [AAA = AMG × 100/(AMG+ALB]. Median PROT, ALB, and AMG (in mg/mmol creatinine) were 20.0, 3.8, and 4.9, respectively. Pathological total proteinuria (>22 mg protein/mmol creatinine) was found in 47% of children (25/53). Only 20% of patients with pathological total proteinuria (5/25) had glomerular proteinuria, whereas 80% (20/25) had tubular proteinuria. Three of five children with glomerular proteinuria had chronic allograft nephropathy. Both AMG and albuminuria negatively correlated with the estimated glomerular filtration rate (eGFR) ( p = 0.021 and 0.003, respectively). In conclusion, tubular proteinuria was present in 80% of children post-RTx and may be associated with impaired graft function; glomerular proteinuria is associated mainly with chronic allograft nephropathy. [ABSTRACT FROM AUTHOR]

BACKGROUND: Mercury is nephrotoxic and dental amalgam is a source of mercury exposure. METHODS: Children 6-10 years of age (n = 534) with two or more posterior teeth with caries but no prior amalgam restorations, were randomized to one of two treatments—amalgam or resin composite (white fillings)—used for caries treatment during 5 years of follow-up. The primary outcome was change in IQ, but important secondary outcomes were effects on markers of glomerular and tubular kidney function: urinary excretion of albumin, alpha-1-microglobulin (A1M), γ-glutamyl transpeptidase (γ-GT), and N-acetyl-β-D-glucosaminidase (NAG). These markers were measured on several occasions during the trial, together with urinary mercury and covariates. We evaluated the results using repeated-measures analyses. RESULTS: There were no significant differences between treatment groups in average levels of renal biomarkers, nor significant effects of number of dental amalgams on these markers. There was, however, a significantly increased prevalence of microalbuminuria (MA) among children in the amalgam group in years 3-5 (adjusted odds ratio 1.8; 95% confidence interval, 1.1-2.9). Most of these cases are likely to be temporary MA, but 10 children in the amalgam group had MA in both years 3 and 5, versus 2 children in the composite group (p = 0.04). There were no differences in the occurrence of high levels of renal tubular markers (A1M, γ-GT, or NAG). CONCLUSIONS: The increase in MA may be a random finding, but should be tested further. The results did not support recent findings in an observational study of an effect of low-level mercury on tubular biomarkers in children. [ABSTRACT FROM AUTHOR]

Abstract: Background: The haem-binding protein α1-microglobulin (α1m) is involved in protection against oxidative damage induced by extracellular haem/haemoglobin. A carboxy-terminally truncated form of α1m (t-α1m), formed by reactions with haemoglobin, degrades haem into a yellow–brown chromophore linked to the protein. The aim of this work was to investigate if t-α1m is present in urine from a large cohort and if urinary and plasma α1m/t-α1m concentrations are changed in patients with haemolytic disorders and haemochromatosis. Methods: Urine and blood from patients (n =20) and a control group (n =22) were investigated for α1m and t-α1m by gel electrophoresis, Western blotting and radioimmunoassay. Data were compared to clinical chemistry data and medical records. Results: Two thirds of all studied subjects displayed t-α1m in urine but the t-α1m/α1m ratio was not increased in patients. Instead, significantly elevated ratios were found in females compared to males. Patients with intravascular or extravascular haemolysis showed higher α1m, albumin and β2-microglobulin/creatinine ratios in urine indicating glomerulo-tubular dysfunction. Conclusions: The demonstration of t-α1m in urine of this cohort may be of importance in quantitative clinical chemistry. Whilst impaired kidney function due to intravascular haemolysis is well-known to occur, it is an unexpected finding in a group of patients with extravascular haemolysis. [Copyright &y& Elsevier]

Absorption, fluorescence, and fluorescence excitation spectra in UV and visible regions are studied for alpha-1-microglobulin preparations isolated from human urine by gel chromatography and immunoaffinity chromatography with charcoal adsorption. The possible nature of low-molecular-weight compounds that impart yellow-brown color to alpha-1-microglobulin preparations and their role in the stabilization of the structure of protein globule is discussed. The effect of urea (1–10 M) and guanidine hydrochloride (0.25–6 M) on the conformational state and fast internal dynamics of alpha-1-microglobulin is studied by tryptophan fluorescence. The unfolding of the protein under the action of denaturants is attended with pronounced activation of its nanosecond internal dynamics. Alpha-1-microglobulin can regain the initial conformation and internal dynamics typical of native protein after denaturation unfolding of the globule with 10 M urea or 6 M guanidine hydrochloride. Alpha-1-microglobulin isolated by gel chromatography can exist in a partially folded thermodynamically stable state in 4–6 M urea. [ABSTRACT FROM AUTHOR]

Predictors of tubular proteinuria (alpha 1-M/crea ratio >10 mg/mmol) were sought in 100 infants of 24–32 weeks’ (group 1) and 69 of 34–42 weeks’ gestation (group 2). Random spot urine samples were obtained in the former group at the ages of 0–3 days, at 1–2 weeks and thereafter at 2-week intervals until the disappearance of tubular proteinuria, and in the latter one sample at a mean (SD) of 3.0 days’ (1.3) age. In group 1, gestational age correlated negatively with the first urinary alpha 1-M/crea ratio. The highest urinary alpha 1-M/crea ratios [median (range) 39.1 mg/mmol (9.5–268.9)] occurred at a median (range) of 5 days’ (1–42) age. Low gestational age and the need for inotropes predicted tubular proteinuria early after birth, whereas low gestation and long duration of ventilator treatment predicted the highest alpha 1-M/crea ratios. Prolonged vancomycin treatment and low gestational age were associated with delayed normalization of tubular proteinuria. In group 2 no significant risk factors for tubular proteinuria were found. The urinary alpha 1-M/crea ratio seems to be a sensitive indicator of renal tubular function in neonates, with low gestational age, the need for inotropes and prolonged assisted ventilation being predictors of increased tubular proteinuria. Long vancomycin courses should be avoided in pre-term infants in view of the prolonged adverse renal effects. [ABSTRACT FROM AUTHOR]