Urinary Total Protein Research Articles

Novel Finding of Urinary Erythropoietin as an Early Biomarker of Cisplatin-Induced Nephrotoxicity.

Cisplatin is a chemotherapeutic drug used to treat a great variety of solid tumors. Its dose is commonly limited by its nephrotoxicity, manifested as acute kidney injury (AKI). Erythropoietin (Epo) is a glycoprotein hormone that regulates the production of red blood cells. This study was performed to evaluate the presence of endogenous Epo in male Wistar rat urine and to analyse changes in urinary Epo levels in response to cisplatin- induced AKI. Dose-dependent studies and time-dependent experiments were performed to evaluate changes in urea nitrogen and creatinine in plasma as well as Epo, neutrophil gelatinase-associated lipocalin (NGAL), alkaline phosphatase (AP) activity, creatinine and total proteins in urine at 2days post-dosing. Rats received 2, 5 or 10mg/kg b.w., i.p. of cisplatin. At 5mg/kg b.w., i.p. cisplatin, significant increases in urinary Epo were detected. Significant increases in urea nitrogen and creatinine in plasma, NGAL, AP, proteins, and Epo were observed in urine from rats that received 10mg/kg b.w., i.p. of cisplatin. In the time-dependent experiments, rats were injected with a dose of 5mg/kg b.w., i.p. of cisplatin, and sampling occurred 2, 4, and 14 days post-dosing. In these animals, there were significant increases in urea nitrogen and creatinine in plasma and total proteins, AP activity, Epo, and NGAL in urine on day 4. Urinary Epo was also detected on day 2. Taken together, these findings provide weight of evidence for urinary Epo as a promising early biomarker of cisplatin-induced AKI in male rats.

Concentration of total proteins in urine as a non-invasive biomarker of endometriosis.

The disease in which we observe the invasion and growth of endometrial cells on extrauterine tissues and organs with the creation of a chronic inflammatory state is called endometriosis. It causes infertility and is present in more than 30% of patients with endometriosis. Diagnosis and treatment of the disease is most often delayed for about 8 years after the first symptoms were reported. The symptomatology of endometriosis is varied, and there is no non-invasive way of diagnosis, and this is the reason for the delayed start of treatment. The development of endometriosis activates pathological processes such as the invasion and proliferation of endometriotic cells, the formation of adhesions and the activation of the immune system, which result in increased protein expression. The aim of this research is to compare the concentrations of total proteins in the urine of subjects with endometriosis with those of the control group and possibly identify a biomarker for the diagnosis of endometriosis. Prospective urine analysis of 141 patients who were hospitalized and surgically treated at the Clinic for Gynecology and Obstetrics of KBC Rijeka from 08/21/2021 until 07/30/2022. The urine of subjects with endometriosis (N=84) and without endometriosis (n=57) was analyzed. Total protein in the urine is increased in the urine of subjects with endometriosis, but the total amount of protein does not correlate with the degree of disease progression. An increase in the level of total proteins in urine in subjects with endometriosis is a possible non-invasive diagnostic biomarker. Patients with endometriosis are grouped after the concentration of total proteins greater than 5000 μg/μl.

Calcineurin B inhibits calcium oxalate crystallization, growth and aggregation via its high calcium-affinity property

Calcineurin inhibitors (CNIs) are widely used in organ transplantation to suppress immunity and prevent allograft rejection. However, some transplant patients receiving CNIs have hypocitraturia, hyperoxaluria and kidney stone with unclear mechanism. We hypothesized that CNIs suppress activities of urinary calcineurin, which may serve as the stone inhibitor. This study aimed to investigate effects of calcineurin B (CNB) on calcium oxalate monohydrate (COM) stone formation. Sequence and structural analyses revealed that CNB contained four EF-hand (Ca2+-binding) domains, which are known to regulate Ca2+ homeostasis and likely to affect COM crystals. Various crystal assays revealed that CNB dramatically inhibited COM crystallization, crystal growth and crystal aggregation. At an equal amount, degrees of its inhibition against crystallization and crystal growth were slightly inferior to total urinary proteins (TUPs) from healthy subjects that are known to strongly inhibit COM stone formation. Surprisingly, its inhibitory effect against crystal aggregation was slightly superior to TUPs. While TUPs dramatically inhibited crystal-cell adhesion, CNB had no effect on this process. Ca2+-affinity assay revealed that CNB strongly bound Ca2+ at a comparable degree as of TUPs. These findings indicate that CNB serves as a novel inhibitor of COM crystallization, growth and aggregation via its high Ca2+-affinity property.

Qing-Re-Xiao-Zheng-Yi-Qi formula relieves kidney damage and activates mitophagy in diabetic kidney disease.

Introduction: Qing-Re-Xiao-Zheng-Yi-Qi Formula is an effective prescription in diabetic kidney disease treatment, we have confirmed the efficacy of Qing-Re-Xiao-Zheng therapy in diabetic kidney disease through clinical trials. In this study, we investigated the mechanisms of Qing-Re-Xiao-Zheng-Yi-Qi Formula in the treatment of diabetic kidney disease. Methods: We used Vanquish UHPLCTM to analyze the chemical profiling of Qing-Re-Xiao-Zheng-Yi-Qi Formula freeze-dried powder. We constructed diabetic kidney disease rat models induced by unilateral nephrectomy and high-dose streptozocin injection. We examined blood urea nitrogen, serum creatinine, serum glucose, total cholesterol, triglyceride, serum total protein, albumin, alanine aminotransferase, aspartate aminotransferase and 24h urinary total protein in diabetic kidney disease rats. The renal pathological changes were observed by HE, Masson, PAS stanning and transmission electron microscopy. The levels of fibrosis-related proteins and mitophagy-related proteins were detected by western blot analysis. We also conducted an immunofluorescence co-localization analysis on podocytes to further investigate the effect of Qing-Re-Xiao-Zheng-Yi-Qi Formula treatment on mitophagy. Results: A total of 27 constituents in Qing-Re-Xiao-Zheng-Yi-Qi Formula were tentatively identified. We found PINK1/Parkin-mediated mitophagy was inhibited in diabetic kidney disease. Qing-Re-Xiao-Zheng-Yi-Qi Formula treatment could raise body weight and reduce renal index, reduce proteinuria, improve glycolipid metabolic disorders, ameliorate renal fibrosis, and reduce the expression of Col Ⅳ and TGF-β1 in diabetic kidney disease rats. Qing-Re-Xiao-Zheng-Yi-Qi Formula treatment could also increase the expression of nephrin, activate mitophagy and protect podocytes in diabetic kidney disease rats and high glucose cultured podocytes. Conclusion: PINK1/Parkin-mediated mitophagy was inhibited in diabetic kidney disease, and Qing-Re-Xiao-Zheng-Yi-Qi Formula treatment could not only ameliorate pathological damage, but also promote mitophagy to protect podocytes in diabetic kidney disease.

Urinary 8-oxoGuo as a potential novel evaluation index for patients with nephrotic syndrome

Urinary 8-oxo-7,8-dihydroguanosine (8-oxoGuo) and 8-oxo-7,8-dihydro-2′- deoxyguanosine (8-oxodGuo) are considered biomarkers of oxidative stress, and patients with nephrotic syndrome have been reported to have increased oxidative stress levels. In this study, we aimed to assess the value of 8-oxoGuo and 8-oxodGuo as novel biomarkers to evaluate the severity of nephrotic syndrome. In total, 107 patients with nephrotic syndrome and 116 healthy controls were recruited for this study. The concentrations of urinary 8-oxoGuo and 8-oxodGuo were measured using isotope-labeled liquid chromatography with tandem mass spectrometry. Urinary creatinine was used to regulate 8-oxoGuo and 8-oxodGuo concentrations. Urinary 8-oxoGuo and 8-oxoGuo/Cr levels in patients with nephrotic syndrome were significantly higher than those in healthy control participants. 8-oxoGuo/Cr showed a positive correlation with the 24 h urinary total protein (UTP) and UTP levels and negative correlations with serum total protein and albumin levels. After treatment, urinary 8-oxoGuo and 8-oxoGuo/Cr levels were significantly lower in the group with a low 24 h-UTP value (<3.5 g/d) than in the high value group. 8-oxoGuo can be used as a feasible and reliable biomarker for the assessment of nephrotic syndrome. Highlights Urinary 8-oxoGuo level was significantly increased in patients with nephrotic syndrome. Urinary 8-oxoGuo level increased with an increase in plasma protein and a decrease in urine protein. Urinary 8-oxoGuo level decreased with nephrotic syndrome remission when urinary microalbumin showed no significant change. Urinary 8-oxoGuo level can be used as novel biomarkers of nephrotic syndrome.

Renal Health Improvement in Diabetes through Microbiome Modulation of the Gut-Kidney Axis with Biotics: A Systematic and Narrative Review of Randomized Controlled Trials.

Diabetes mellitus is the most common endocrine disorder worldwide, with over 20% of patients ultimately developing diabetic kidney disease (DKD), a complex nephropathic complication that is a leading cause of end-stage renal disease. Various clinical trials have utilized probiotics, prebiotics, and synbiotics to attempt to positively modulate the gut microbiome via the gut-kidney axis, but consensus is limited. We conducted a multi-database systematic review to investigate the effect of probiotics, prebiotics, and synbiotics on various biomarkers of renal health in diabetes, based on studies published through 10 April 2022. Adhering to the Cochrane Collaboration and Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, relevant articles were systematically screened and extracted by independent reviewers; subsequently, results were systematically compiled, analyzed, and expanded through a narrative discussion. A total of 16 publications encompassing 903 diabetic individuals met the inclusion criteria. Our findings show that some studies report statistically significant changes in common renal markers, such as serum creatinine, estimated glomerular filtration rate, blood urea nitrogen/urea, microalbuminuria, and uric acid, but not on serum albumin, sodium, potassium, phosphorous, or total urine protein. Interestingly, these nutraceuticals seem to increase serum uric acid concentrations, an inflammatory marker usually associated with decreased renal health. We found that probiotics from the Lactobacillus and Bifidobacterium families were the most investigated, followed by Streptococcus thermophilus. Prebiotics including inulin, galacto-oligosaccharide, and resistant dextrin were also examined. The single-species probiotic soymilk formulation of Lactobacillus plantarum A7 possessed effects on multiple renal biomarkers in DKD patients without adverse events. We further investigated the optimum nutraceutical formulation, discussed findings from prior studies, described the gut-kidney axis in diabetes and DKD, and finally commented on some possible mechanisms of action of these nutraceuticals on renal health in diabetics. Although probiotics, prebiotics, and synbiotics have shown some potential in ameliorating renal health degradation in diabetes via gut-kidney axis crosstalk, larger and more convincing trials with focused objectives and next-generation nutraceutical formulations are required to investigate their possible role as adjunct therapy in such patients.

Evaluation of Novel Urinary Biomarkers in Beagle Dogs With Amphotericin B-Induced Kidney Injury.

Next-generation urinary protein biomarkers have been qualified to enable monitoring for drug-induced kidney injury in toxicology studies conducted in rats. However, there is limited literature on the utility of these biomarkers in dogs. To add to the existing body of knowledge on the utility of the next-generation drug-induced kidney injury (DIKI) biomarkers, we evaluated the value of these biomarkers for the early detection of DIKI in Beagle dogs using a differentiated nephrotoxicant, Amphotericin B (AmpB). In dogs with AmpB-induced kidney injury, we monitored the response of urinary albumin, total protein, clusterin, kidney injury molecule 1, neutrophil gelatinase-associated lipocalin and N-acetyl-beta-D-glucosaminidase. We also measured blood urea nitrogen, serum creatinine and cystatin C. The results showed that urinary clusterin (up to ∼ 112x) was much more sensitive to AmpB-induced kidney injury relative to other biomarkers. Moreover, other than urinary clusterin and to a much lesser extent urinary albumin and total protein, none of the other biomarkers analyzed in this study were more sensitive than blood urea nitrogen and serum creatinine. The AmpB related tubular alterations were characterized by minimal to mild, multifocal necrosis, degeneration, regeneration, dilatation and mineralization. The mild nature of these histopathologic findings further attested to the sensitivity of urinary clusterin to AmpB-induced kidney injury in dogs. These results will help drug developers make informed decisions when selecting urinary biomarkers for monitoring DIKI in dogs for toxicology studies.

ODP242 Safety of SGLT2-Inhibitors in Patients with Multiple Myeloma and Diabetes

Abstract Background An estimated 10-20% of patients with multiple myeloma (MM) have type 2 diabetes (T2DM). About half of patients with MM will experience renal insufficiency,the risk of which increases in the setting of T2DM. While sodium-glucose cotransporter 2 inhibitors (SGLT2i) are beneficial in patients with chronic kidney disease (CKD) due to diabetic or hypertensive nephropathy, the role of SGLT2i in MM-related CKD is unknown. Moreover, as MM therapies target the immune system, concurrent use with SGLT2i may increase the risk of adverse events such as infections. The current work is the first to examine the potential benefit and safety of SGLT2i in patients with MM and T2DM. Methods This study was approved by the Institutional Review Board. A retrospective cohort study was performed on patients aged 18 years or older with MM who received SGLT2i therapy between March 2013 and December 2020 at a quaternary academic medical center. Electronic medical records were reviewed for the following data: demographics, comorbidities, MM parameters, medication and medical history, SGLT2i usage and duration, and hemoglobin A1c(HbA1c) and serum creatinine pre- and post-SGLT2i initiation. Data are presented as mean ± standard deviation or counts and percentages, and was analyzed using SPSS software (version 25. 0, IBM Corp). Results In total, 50 patients were identified. Patients who were diagnosed with MM after initiation of SGLT2i (n= 12), did not have any MM data available (n=1), or for whom SGLT2i usage could not be verified (n= 4) were excluded. Thirty-three patients were included in the final analysis. The mean age was 63 (±7.8) years, 63.6% were male, and 87.9% had active MM or MM in remission. Most had received stem-cell transplantation (55. 0%) and multiple lines of chemotherapy (63.6%) prior to SGLT2i initiation. The average HbA1c was 7.4% (±1.2%) and 20% of patients had CKD prior to SGLT2i initiation. The overall SGLT2i discontinuation rate was 42.4%, and the mean duration of therapy was 2.2(±1.5) years. Notably, 21.4% of discontinuation events were due to an increase in creatinine; however, for the entire cohort, there were no statistically significant changes in serum creatinine, total body weight, HbA1c, or 24-hour urine protein at 15 months’ follow-up. Other reasons for SLGT2i discontinuation included loss to follow-up, change of health insurance coverage for SGLT2i, and unrelated hospital admission. Genitourinary infection was not cited as a reason for any SGLT2i discontinuation. Conclusion SGLT2i therapy may be safe in patients with MM and T2DM, but no benefit was found with respect to serum creatinine, HbA1c, or 24-hour urine protein. Prospective research is required to further evaluate the potential benefit and safety of SGLT2i therapy in this population. Presentation: No date and time listed

Efficacy of chlorogenic acid against ethylene glycol-induced renal stone model: The role of NFKB-RUNX2-AP1-OSTERIX signaling pathway

Background and objectiveRenal tissue injuries by free radicals are an essential reason in pathogenesis of urinary tract stones. Ethylene glycol is one of the toxic agents which can causes to the increases in biosynthesis of reactive oxygen species and oxidative stress condition. Natural antioxidants have been reported to protective efficacy against renal stones formation. Accordingly, the aim of the current experiment was to identify the renal protective effect of chlorogenic acid as a well-prominent antioxidant on ethylene glycol-induced renal stone model targeting the NFKB-RUNX2-AP1-OSTERIX signaling pathway. Materials and methodsRenal stones model were established by ethylene glycol (Percent: 0.75) within the daily drinking water for rats. Treatment groups received cystone (750 mg/kg) and chlorogenic acid (100, 200, and 400 mg/kg, day: 15th to 28th, gavage). After 4 weeks, the renal function parameters (calcium, uric acid, creatinine, total protein, oxalate, and citrate) in plasma, urine, and renal tissue were measured. Moreover, oxidative stress factors and gene expression of NFKB, RUNX2, AP1, and OSTERIX were also evaluated. ResultsThe results showed improved renal function in chlorogenic acid-treated groups. The total proteins and creatinine excretion were decreased. Also the gene expression of oxidative stress pathway (NFKB-RUNX2-AP1-OSTERIX) were decreased which caused to increases of antioxidant enzymes. Conclusionsthe antioxidant activity increases by chlorogenic acid treatment may have a critical role in prevention of calcium oxalate formation via inhibition of the NFKB-RUNX2-AP1-OSTERIX signaling pathway.

Urine stabilization and normalization strategies favor unbiased analysis of urinary EV content

Urine features an ideal source of non-invasive diagnostic markers. Some intrinsic and methodological issues still pose barriers to its full potential as liquid biopsy substrate. Unlike blood, urine concentration varies with nutrition, hydration and environmental factors. Urine is enriched with EVs from urinary-genital tract, while its conservation, purification and normalization can introduce bias in analysis of EV subsets in inter-and intra-individual comparisons. The present study evaluated the methods that decrease such biases such as appropriate and feasible urine storage, optimal single-step EV purification method for recovery of proteins and RNAs from small urine volumes and a normalization method for quantitative analysis of urine EV RNAs. Ultracentrifugation, chemical precipitation and immuno-affinity were used to isolate EVs from healthy donors’ urine that was stored frozen or at room temperature for up to 6 months. Multiple urine biochemical and EV parameters, including particle count and protein content, were compared across urine samples. To this purpose nanoparticle tracking analysis (NTA) and protein assessment by BCA, ELISA and WB assays were performed. These measurements were correlated with relative abundances of selected EV mRNAs and miRNAs assessed by RT-PCR and ranked for the ability to reflect and correct for EV content variations in longitudinal urine samples. All purification methods enabled recovery and downstream analysis of EVs from as few as 1 ml of urine. Our findings highlight long term stability of EV RNAs upon urine storage at RT as well as excellent correlation of EV content in urine with some routinely measured biochemical features, such as total urine protein and albumin, but not creatinine most conventionally used for urine normalization. Comparative evaluation of mRNA and miRNAs in EV isolates revealed specific RNAs, in particular RNY4 and small miRNA panel, levels of which well reflected the inter-sample EV variation and therefore useful as possible post-analytical normalizers of EV RNA content. We describe some realistic urine processing and normalization solutions for unbiased readout of EV biomarker studies and routine clinical sampling and diagnostics providing the input for design of larger validation studies employing urine EVs as biomarkers for particular conditions and diseases.

Urinary RBP as an Independent Predictor of Renal Outcome in Diabetic Nephropathy.

Background Renal tubular impairment is prevalent in diabetic nephropathy (DN) and the histological severity predicted renal outcome. Biomarkers of tubular injury also increased in the urine of DN patients. The retrospective study aimed to assess the prognostic value of clinically widely applied urinary tubular injury markers, retinol-binding protein (RBP), β2-microglobulin (β2-MG) and N-acetyl-β-D-glucosaminidase (NAG) in DN. Method A total of 305 patients with biopsy-proven DN were enrolled. The baseline urine total protein and components including albumin, IgG, RBP, β2-MG and NAG were retrieved from medical records. The primary outcome was end stage renal disease (ESRD). Cox proportional hazard analysis and restricted cubic splines were performed to evaluate the association of parameters with ESRD. Nomograms were constructed and concordance index (C-index) was used to measure the prediction ability. Result The levels of urinary RBP, β2-MG and NAG were positively correlated with the severity of interstitial fibrosis and tubular atrophy (IFTA). Positive correlations were also observed among β2-MG, NAG and mesangial expansion. Urinary RBP was not correlated with any glomerular lesions. Urinary RBP, β2-MG and NAG were risk factors for ESRD in hazard analysis with adjustment for age, gender and body mass index (BMI). The hazard ratios increased with the increment of baseline levels. In the multivariate Cox model including serum creatinine (SCr), total urinary protein, urinary albumin, urinary IgG and the tubular injury biomarkers, urinary RBP (with every g/mol.Cr increase: HR 1.06, 95% CI 1.03-1.10, p =0.001) remained as an independent risk factor for ESRD in DN patients. Patients were divided by the medium value of urinary RBP into the low RBP and high RBP groups. Survival analysis showed that significantly more patients in the high RBP progressed to ESRD compared to those in the low RBP group (p =0.02) when urinary total protein was less than 3.5 g/g. The C-index of the nomogram incorporating age, gender, BMI, SCr and total urine protein was 0.757. The value increased to 0.777 after adding urinary RBP to the model. Conclusions Urinary RBP excretion was only correlated with the severity of IFTA and independently predicted ESRD in DN patients.

Anti-urolithiatic Activity of Daidzin in Ethylene Glycol-Induced Urolithiasis in Rats.

Urolithiasis is a common urological disorder, which causes considerable morbidity in both genders at all age groups worldwide. Though treatment options such as diuretics and non-invasive techniques to disintegrate the deposits are available, but often they are found less effective in the clinics. In this work, we planned to investigate the ameliorative effects of daidzin against the ethylene glycol (EG)-induced urolithiasis in rats. The male albino rats were distributed into four groups (n = 6) as control (group I), urolithiasis induced by the administration of 0.75% EG (group II), urolithiasis induced rats treated with 50mg/kg of daidzin (group III), and urolithiasis rats treated with standard drug 750mg/kg of cystone (group IV). The urine volume, pH, and total protein in the urine were assessed. The activities of marker enzymes in both plasma and kidney tissues were analyzed using assay kits. The levels of kidney function markers such as calcium, oxalate, urea, creatinine, uric acid, magnesium, BUN, and phosphorous were estimated using assay kits. The status of antioxidants and inflammatory cytokines were also examined using kits. The renal tissues were examined by histopathological analysis. Our results revealed that the daidzin treatment effectively decreased the urine pH and protein level and increased the urine volume in the urolithiasis rats. Daidzin decreased the calcium, oxalate, uric acid, and urea, creatinine, and BUN levels and also improved the magnesium and phosphorus in the urolithiasis rats. The activities of AST, ALT, ALP, GGT, and LDH were effectively reduced by the daidzin in both serum and renal tissue. Daidzin also reduced the inflammatory marker and increased the antioxidant levels. Histopathology results also proved the therapeutic effects of daidzin. Together, our results displayed that daidzin is effective in the amelioration of EG-induced urolithiasis in rats.

Effects of Shenkang Injection Combined with Jinshuibao on Early Diabetic Nephropathy and Effects on Coagulation Fibrinolysis System and Urinary Protein.

Purpose To explore the effect of Shenkang injection (SKI) combined with Jinshuibao for early diabetic nephropathy (DN) and its effect on the coagulation fibrinolysis system and urinary protein. Methods 136 patients with early DN admitted to our hospital from March 2018 to October 2019 were divided into the observation group (n = 68) and the control group (n = 68) randomly. On the basis of the conventional treatment, the control group was treated with SKI, and the observation group was treated with SKI and Jinshuibao. Two weeks later, the therapeutic effects of the 2 groups were compared. The prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT), fibrinogen (FIB), tissue plasminogen activator (t-PA), plasminogen activator inhibitor-1 (PAI-1), and D-dimer (D-D) were observed and compared before and after the treatment. 24 hour urine total protein (24 h-UTP), urine albumin excretion rate (UAER), and urine β2 microglobulin (β2-MG) were measured and compared before and after the treatment. Adverse reactions in the two groups were recorded during the treatment. Results The effective rate of the observation group after treatment was 92.65% higher than the control group 79.41%. the difference was statistically significant (P < 0.05). The levels of PT, APTT, TT, FIB, PAI-1, and D-D in the two groups after treatment were lower, and t-PA levels after treatment were higher than those before, and all of the above indicators were significantly changed in the observation group than in the control group. The difference was statistically significant (P < 0.05). The 24 h-UTP, UAER, and β2-MG in the two groups after treatment were lower than those before, and all of the above indicators were significantly changed in the observation group than in the control group. The difference was statistically significant (P < 0.05). There was no statistically significant difference during the treatment for 2 groups in terms of adverse reactions. The difference was statistically significant (P > 0.05). Conclusion SKI combined with Jinshuibao has a significant effect in the treatment of early DN, which can reduce the risk of hyperfunction of coagulation and fibrinolysis system, further reduce the content of urine protein, and delay the process of DN.

The effect of alcoholic extract of Nerium oleander leafs on some biochemical parameters and function of liver and kidney in white rats

The present study was conducted to know the effect of alcoholic extract of Nerium oleander leafs on some biochemical parameters as well as the histological changes in liver and kidney in male albino rats. Twenty mature albino rats were randomly divided one word groups: control group and treatment group. Control group was treated with drinking water for (6 weeks), while treatment group was treated with 4 mg/kg B.W of alcoholic extract of Nerium oleander leafs. Blood samples were collected every 3 weeks to make biochemical parameters. The results showed significant decrease (P&lt;0.05) in Total protein , albumin and significant increase (P&lt;0.05) in amino transmission enzymes concentration (AST and ALT) , urine and keratinin concentration in treatment group compared with control group. The study reveals high toxic effect of plant on all body tissues.

Histopathologic and clinical features of diabetic nephropathy alone and with concomitant nondiabetic renal diseases

Background and objective: Diabetic nephropathy (DN) is a major complication of diabetes mellitus (DM) and one of the leading causes of end-stage kidney disease. The aim of the present study was to evaluate the histomorphological and clinical profiles of DN and associated non-diabetic renal dieases (NDRD) in diabetic patients. Materials and methods: The study was carried out at the Department of Histopathology, Armed Forces Institute of Pathology (AFIP), Dhaka, from July 2019 to December 2020. Renal biopsy samples from known diabetic patients were included in the study. The formalin-fixed tissues were stained with haematoxylene &amp; eosin (H&amp;E), Periodic acid Schiff (PAS), Masson Trichrome (MT) and Jones Methanamine Silver (JMS) stains. Tissues were stained for IgG, IgA, IgM, C3, C1q, kappa and lambda for direct immunofluorescence (DIF) study. DN was histologically classified according to Tervaert classification system. Interstitial fibrosis and tubular atrophy (IFTA) as well as arteriolar hyalinization scoring was also done. Clinical information was retrieved from the patient’s information sheet. Results: Total 46 biopsy samples from DN cases were included in the study. The mean age of the cases was 46.76+10.63 years, including 36 males and 10 females. The most common clinical presentation was nephritic range proteinuria (n=32, 69.56%). Among all, 27 (58.69%) patients had haematuria. The mean serum creatinine level was 4.28+2.61 mg/dl, and 80.43% had serum creatinine levels &gt;1.5 mg/dl. Histopathologic examinatiom revealed type III DN in 26 (56.5%) and type IV DN in 11 (23.9%) cases. IFTA score 1 (&lt;25%) was seen in 20 (43.5%), score 2 (25-50%) in 19 (41.3%) and score 3 (&gt;50%) in 7 (15.2%). Vascular hyalinization score-2 in 25 (54.3%), score-1 in 14 (30.4%) and score-0 in 7 (15.2%). DN class II, III and IV were associated with high urinary total protein (UTP) and serum creatinine levels. Among the histologic changes, percentage of glomerular sclerosis, the mean IFTA score and vascular hyalinization score were found to be highest in class IV DN, and all were significantly associated with histologic glomerular DN classes (p= &lt;0.05). Of the total cases, 21 (45.65%) were found with nondiabetic renal diseases (NDRD), the most common feature was focal segmental glomerulosclerosis (FSGS) (26.57%), followed by IgA nephropathy and post-infectious glomerulonephritis (PIGN). Among 46 cases, one post-transplant biopsy was included, which revealed class II DN along with features of calcineurin inhibitor toxicity. Conclusion: Tervaert’s histologic classification of our cases revealed class III DN lesions as the predominant one, and the classes had a significant association with age of the patient, serum creatinine level, mean IFTA, arteriolar hyalinization and NDRD. Among the NDRD, FSGS was the most common pathology. IMC J Med Sci. 2023; 17(1): 003. DOI: https://doi.org/10.55010/imcjms.17.003 *Correspondence: Sk Md Jaynul Islam, Department of Histopathology, Armed Forces Institute of Pathology, Dhaka Cantonment, Dhaka, Bangladesh. Email: jaynul.islam@gmail.com

P38 The gut that leaks

Introduction/BackgroundSystemic lupus erythematosus is a chronic inflammatory autoimmune disease which can affect multiple organ systems. Symptoms of intestinal involvement in SLE may vary from common and non-specific such as heartburn, oral ulcers and dysphagia to less common but more serious including abdominal pain due to serositis and intestinal vasculitis.Description/MethodA 20 year old girl was referred to the gastroenterology team for the management of ascites. Her past medical history included SLE diagnosed aged 10, class IV lupus nephritis, autoimmune haemolytic anaemia secondary to SLE, Haemoglobin H, gastrostomy feeding due to poor growth.At the time of the transition from paediatric to adult service her SLE was under good control. She was on Prednisolone 6 mg on alternate days, Mycophenolate (MMF) 750 mg BD, Septrin 240 mg OD and Enalapril 2.5 mg OD. After the transition, MMF was reduced to 500 mg OD. The rest were unchanged. Serum total protein was 57 (60-80), serum albumin 32 (35-50). Weight was 41.2 kg.Ten months after the transition, she presented with bilateral leg swelling and subsequently tense ascites over 3 weeks. Serum total protein reduced to 31, albumin 13.24 hour urine total protein was 0.15 (0-0.1), urine volume 1.7 L. Serum creatinine 20 µmol/L and eGFR >90. Urine PCR was 16 mg/mmol.Total protein in ascites drain fluid was <20 g/L, albumin <10 g/L, glucose 5.7 mol/L. There were no malignant cells and no growth on enrichment culture.She was referred to the dietitians who advised that her oral intake was adequate.There was no evidence of heart failure on echocardiogram.Small bowel meal showed oedematous bowel wall but no abnormalities in terminal ileum. Coeliac serology was negative.CT abdomen and pelvis showed normal appearances to the liver, hepatic and portal veins; no features of Budd-Chiari syndrome; normal size kidneys; incidental finding of non-occlusive right internal jugular thrombus and moderate lymphadenopathy in mesenteric and retroperitoneal regions.She developed swelling in the right calf. Doppler ultrasound confirmed major DVT with occlusive thrombus extending to distal superficial femoral veins. Clots in inferior vena cava and portal veins were excluded.Discussion/ResultsDuodenal biopsy confirmed villous oedema and dilated lymphatics consistent with changes seen in protein losing enteropathy. She was referred to Rheumatology. Prednisolone was increased to 15 mg. The response was rapid with reduction in ascites. MMF was increased to maximum tolerable dose 750 mg AM and 500 mg PM.Hb had been persistently low around 50 during this illness. She had longstanding hypocomplementaemia. C3 dropped to 0.37 (baseline 0.6). C4 remained low around 0.04. IgG was 5.9 (baseline >7). Serology showed DsDNA was 5 (normal range 0-15), positive ANA, trace positive RNP and negative Rheumatoid factor and APL antibodies. CRP was 10.Abdominal lymphadenopathy was felt to be due reactive changes due to active lupus.With increased steroids and immunosuppressants, she improved significantly both clinically and biochemically within a few weeks. Hb rose to over 90, serum albumin > 25 and serum protein above 50 by the end of 3 months after treatment escalation.Warfarin was stopped upon completion of 6 month anticoagulation.On repeat CT, previous abdominal lymphadenopathy had resolved completely. Splenomegaly measuring 17.6 cm was noted. This was discussed in haematology MDT and thought to be due to the haemoglobinopathy and haemolysis.Haemolysis continued to be a challenge. She remained on relatively high dose steroids varying between 20 mg and 10 mg over the years. Vitamin D was prescribed. Weight was very low but stable just above 38 kg.Protein Losing Enteropathy (PLE) is rare but recognised in SLE. It typically occurs in young women with clinically severe disease with multi organ involvement. Clinical presentation is characterised by the onset of profound oedema, hypoalbuminaemia in the absence of nephrotic range proteinuria. Severe diarrhoea is present in 50% of the patients. It is important to investigate and exclude other causes of hypoalbuminaemia (e.g cardiac failure, liver diseases, malnutrition) and PLE.Key learning points/ConclusionPLE can be the initial manifestation of SLE.Lupus nephritis and PLE can be present successively or simultaneously in a lupus patient.Stringent diagnostic evaluation is required to exclude other mimicking conditions.Multidisciplinary team involvement is crucial in the management of patients with multi-organ involvement.PLE associated with SLE usually responds well to corticosteroids. However additional immunosuppressant(s) are generally required to treat other SLE symptoms as well as to help with the steroid reduction regime.As well as treating the underlying disease, advice from dietitians is important to improve and maximise the nutritional status of these patients.A clinically significant hyper-coagulable state is reportedly attributed to PLE associated with SLE. Therefore, VTE prophylaxis especially in hospitalised patients is recommended. Moreover, clinicians should have high index of suspicion for VTE in these patients with multiple risk factors to arrange early appropriate investigations and treatment plan.

Cystathionine β-synthase gene inactivation dysregulates major urinary protein biogenesis and impairs sexual signaling in mice.

Reproductive success in mice depends on sexually dimorphic major urinary proteins (Mup) that facilitate interactions between females and males. Deletion of cystathionine β-synthase (Cbs) gene, a metabolic gene important for homeostasis of one-carbon metabolism, impairs reproduction by causing female infertility in mice. Here, we examined Mup biogenesis and sexual signaling in Cbs-/- versus Cbs+/- mice. We found significantly reduced levels of total urinary Mup protein in male and female Cbs-/- versus Cbs+/- mice. SDS-PAGE/Western blot, ESI-MS, and RT-qPCR analyses of the liver, plasma, and urinary proteins identified a male-specific Mup20 in Cbs-/- , but not in Cbs+/- females. The 18 893 Da Mup20 became the most abundant in urine of Cbs-/- females and males. Effects of Cbs genotype on 18 645 Da, 18 693 Da, and 18 709 Da Mup species abundance were Mup- and sex-specific. Cbs genotype-dependent changes in hepatic Mups and Mup20 expression were similar at the protein and mRNA level. Changes in Mups, but not in Mup20, can be explained by downregulation of hepatic Zhx2 and Ghr receptors in Cbs-/- mice. Behavioral testing showed that Cbs+/- females ignored Cbs-/- male urine but were attracted to Cbs+/- male urine. Cbs+/- males ignored urine of Cbs-/- males but countermarked urine of other Cbs+/- males and were attracted to urines of Cbs-/- as well as Cbs+/- females. Cbs-/- males did not countermark urine of Cbs+/- males but were attracted to urines of Cbs+/- females. Taken together, these findings show that Cbs, a metabolic gene, interacts with the processes involved in Mup biogenesis that are essential for the maintenance of sexual dimorphism and signaling and suggest that dysregulation of these interactions impairs reproductive fitness in mice.

240.3: Detection of CMV Mediated Sub-Clinical Rejection by Urine Biomarkers

Methods: Research on early detection of Cytomegalovirus (CMV) mediated sub-clinical rejection (sAR) is in its nascence. This study explores the efficacy of urine-based biomarkers for sAR risk prognostication. Urine samples were serially collected (1 week- 2yrs) across 125 unique renal allograft recipients from UCSF and UCLA. These CMV+ (blood CMV-PCR) patients received valganciclovir therapy until CMV- for 3 consecutive months. Analysis: We computed a machine-learning based rejection prediction score incorporating urine cell-free DNA (cfDNA), methylated cfDNA, total protein, CXCL10, clusterin and creatinine. Rejection was determined between scores of 32-100; higher score associated with biopsy severity (Yang et al, STM 2021). In a cross-sectional analysis, Kruskal-Wallis test was performed on the urine-score distribution, to determine sAR during CMV+. Subsequently UCSF samples were analyzed longitudinally to establish the relationship between CMV and sAR; this was validated independently in UCLA samples. Results: Significant differences were observed in scores >32 between biopsy-confirmed AR vs. stable (p: 0.0034) and CMV vs. stable (p=0.003). Longitudinal analysis yielded 3 primary sAR patterns following CMV infection; (i)persistence of sAR leading to new development of chronic allograft injury; (ii)early sAR (score >32) followed by a further increase in score (>55) with biopsy confirmed cAR; (iii)sAR (>32) followed by recovery of injury with CMV resolution and a decline in score <32). Conclusions: We provide a urine-based method for detecting sAR at the time of CMV infection, and its potential clinical utility for titrating use of anti-viral therapy and recipient immunosuppression load.

NMRpQuant: an automated software for large scale urinary total protein quantification by one-dimensional 1H NMR profiles.

Summary 1H nuclear magnetic resonance (NMR) spectroscopy is an established bioanalytical technology for metabolic profiling of biofluids in both clinical and large-scale population screening applications. Recently, urinary protein quantification has been demonstrated using the same 1D 1H NMR experimental data captured for metabolic profiling. Here, we introduce NMRpQuant, a freely available platform that builds on these findings with both novel and further optimized computational NMR approaches for rigorous, automated protein urine quantification. The results are validated by interlaboratory comparisons, demonstrating agreement with clinical/biochemical methodologies, pointing at a ready-to-use tool for routine protein urinalyses.Availability and implementation NMRpQuant was developed on MATLAB programming environment. Source code and Windows/macOS compiled applications are available at https://github.com/pantakis/NMRpQuant, and working examples are available at https://doi.org/10.6084/m9.figshare.18737189.v1.Supplementary information Supplementary data are available at Bioinformatics online.

Physalis angulata leaf extract ameliorates L-NG-nitroarginine methyl ester (L-NAME)-induced preeclampsia symptoms in rats through improved endothelial progenitor cells and endothelial cells due to reduced antiangiogenic factor and oxidative stress

Background: This study aimed to determine the ameliorative effects of Physalis angulata leaf extract on L-NG-nitroarginine methyl ester (L-NAME)-induced preeclampsia symptoms in rats. Methods: Phytochemical analysis of the extract was performed with liquid chromatography-high resolution mass spectrometry (LC-HRMS). Pregnant Wistar rats were randomly divided into five groups (n=6). Preeclampsia rats were injected with L-NAME on gestation days 9 to 18 (G9–G18), while sham rats were injected with the same vehicle volume. Three groups of preeclampsia rats were orally supplemented with the extract at doses of 500, 1500, and 2500 mg/kg body weight/day on G12–G18. The tail-cuff method measured blood pressures. Spectrophotometry determined urine protein levels. Serum soluble fms-like tyrosine kinase (sFlt)-1 levels were evaluated using an enzyme-linked immunosorbent assay (ELISA). Serum and placental malondialdehyde (MDA) levels, superoxide dismutase (SOD) activities, and nitric oxide (NO) levels were measured by colorimetry. Immunohistochemistry was used to determine tail artery endothelial nitric oxide synthase (eNOS), placental eNOS, and placental hypoxia-inducible factor (HIF)-1α expressions. Circulating angiogenic cells (CACs) and endothelial colony-forming cells (ECFCs) were counted using flow cytometry. Results: L-NAME injection increased blood pressures, 24-h total urine protein level, serum sFlt-1 level, serum and placental MDA levels, percentages of CACs and ECFCs, and placental HIF-1α expression. It also decreased serum and placental SOD activities, serum NO level, tail artery and placental eNOS expressions compared to the sham group. Physalis angulata leaf extract administration lowered blood pressures, urine protein level, sFlt-1 level, MDA levels, percentages of CACs and ECFCs, and placental HIF-1α expression. The extract increased SOD activities, NO level, tail artery and placental eNOS expressions compared to the preeclampsia group. Conclusions: Physalis angulata leaf extract reduces antiangiogenic factor and oxidative stress. It also enhances eNOS/NO signaling. Thus, it improves EPC and endothelial cell function and reverses L-NAME-induced hypertension and proteinuria in preeclampsia rats.