Safety and efficacy of Empagliflozin in Pakistani Muslim patients with type 2 diabetes (SAFE-PAK); a randomized clinical trial.

BACKGROUND: Sodium-Glucose-Co-Transporter 2 (SGLT2) inhibitor (Empagliflozin) is an effective drug in controlling blood glucose through predominantly glycosuria. Glycosuria increases the risk of genitourinary infections in diabetes. This study was aimed to establish the safety and efficacy of Empagliflozin (Group-A) versus standard care (Group-B) in Pakistani Muslim individuals with type 2 diabetes. METHODS: A multicenter, randomized clinical trial was conducted in five cities across Pakistan from July 2019 to August 2020. Patients of both genders aged 18-75 years, body mass index (BMI) ≤ 45 kg/m2, glycosylated hemoglobin (HbA1c) 7-10% (53 mmol/mol to 86 mmol/mol) and treatment-naive to Empagliflozin were included. Treatment was given for 24 weeks, and allocation was done through randomization. RESULTS: Out of 745 screened patients, 333 met the eligibility criteria, and a total of 244 (73.3%) patients were enrolled. More hypoglycemic events were reported in the standard care group, whereas positive urine culture, fungal infection, dehydration, and hypotension occurrence were comparable between the two groups. The 6 months mean HbA1c reduction was significant in both groups; (Group-A: 0.91 ± 0.15; p < 0.001 vs. Group-B2: 0.79 ± 0.14; p < 0.001). Efficacy comparison at 6 months revealed a significant reduction in weight and systolic blood pressure (SBP) in Group A only (Group-A: 1.4 ± 0.4 kg; p < 0.002 vs. Group-B: 0.01 ± 0.5 kg; p < 1.00), (Group-A: 5.1 ± 1.7 mmHg; p < 0.012 vs. Group-B: 2.3 ± 1.7 mmHg; p < 0.526). CONCLUSIONS: Empagliflozin was a safe drug compared to standard care in Pakistani Muslim patients with diabetes. It was as effective as standard care in the clinical setting but achieved glycemic control by reducing weight and SBP in type 2 diabetes patients. TRIAL REGISTRATION: This study was registered in the NIH US National Library of Medicine clinical trials registry at Clinicaltrials.gov with the registration number: NCT04665284 on 11/12/2020.

PodoSighter: A Cloud-Based Tool for Label-Free Podocyte Detection in Kidney Whole-Slide Images.

BACKGROUND: Podocyte depletion precedes progressive glomerular damage in several kidney diseases. However, the current standard of visual detection and quantification of podocyte nuclei from brightfield microscopy images is laborious and imprecise. METHODS: We have developed PodoSighter, an online cloud-based tool, to automatically identify and quantify podocyte nuclei from giga-pixel brightfield whole-slide images (WSIs) using deep learning. Ground-truth to train the tool used immunohistochemically or immunofluorescence-labeled images from a multi-institutional cohort of 122 histologic sections from mouse, rat, and human kidneys. To demonstrate the generalizability of our tool in investigating podocyte loss in clinically relevant samples, we tested it in rodent models of glomerular diseases, including diabetic kidney disease, crescentic GN, and dose-dependent direct podocyte toxicity and depletion, and in human biopsies from steroid-resistant nephrotic syndrome and from human autopsy tissues. RESULTS: The optimal model yielded high sensitivity/specificity of 0.80/0.80, 0.81/0.86, and 0.80/0.91, in mouse, rat, and human images, respectively, from periodic acid-Schiff-stained WSIs. Furthermore, the podocyte nuclear morphometrics extracted using PodoSighter were informative in identifying diseased glomeruli. We have made PodoSighter freely available to the general public as turnkey plugins in a cloud-based web application for end users. CONCLUSIONS: Our study demonstrates an automated computational approach to detect and quantify podocyte nuclei in standard histologically stained WSIs, facilitating podocyte research, and enabling possible future clinical applications.

Asymptomatic urinary tract infections and associated risk factors in Pakistani Muslim type 2 diabetic patients.

BACKGROUND: One of the leading long-term complications of type 2 diabetes mellitus (T2DM) includes renal dysfunction and urinary tract infections (UTI) which are considered to be prevalent in uncontrolled diabetes. Moreover, physiological factors like age, gender, duration of diabetes, other diabetic complications like neuropathy, autonomic neuropathy and glycosuria are also considered as predisposing factors for increased prevalence of UTI in diabetes which can be symptomatic or asymptomatic. METHODS: This was a cross-sectional, multi-centre study including diabetic patients from 12 clinical sites spread across major cities of Pakistan. The inclusion criteria were adult Pakistani population of age between 18 to 75 years both genders and suffering from T2DM irrespective of duration. A detailed clinical history of the past 3 months was recorded and, biochemical investigations of blood samples were conducted. Urine culture analysis performed identified the type of pathogen present and was done only for asymptomatic patients. RESULTS: A total of 745 type 2 diabetic patients were initially screened, out of 545 patients considered for final analysis 501 (91.92%) were negative and the rest 44 (8.08%) had positive urine culture. Female gender had a significantly higher proportion of positive urine culture (77.27%, p-value< 0.001). Body mass index and mean age had insignificant distribution among the two groups of positive and negative urine culture, with age 40-59 years having higher proportion (70.45%) in the positive group. Escherichia coli was detected in most of the positive samples (52.3%). All bacterial samples were found resistant to Ciprofloxacin. CONCLUSION: Diabetic Pakistani muslim female patients are identified to be at high risk of suffering from asymptomatic UTI and age more than 40 years is an important risk factor. Escherichia coli was the most common causative organism among people living in this geographical area.

Symptom-based patient stratification in mental illness using clinical notes.

Mental illnesses are highly heterogeneous with diagnoses based on symptoms that are generally qualitative, subjective, and documented in free text clinical notes rather than as structured data. Moreover, there exists significant variation in symptoms within diagnostic categories as well as substantial overlap in symptoms between diagnostic categories. These factors pose extra challenges for phenotyping patients with mental illness, a task that has proven challenging even for seemingly well characterized diseases. The ability to identify more homogeneous patient groups could both increase our ability to apply a precision medicine approach to psychiatric disorders and enable elucidation of underlying biological mechanism of pathology. We describe a novel approach to deep phenotyping in mental illness in which contextual term extraction is used to identify constellations of symptoms in a cohort of patients diagnosed with schizophrenia and related disorders. We applied topic modeling and dimensionality reduction to identify similar groups of patients and evaluate the resulting clusters through visualization and interrogation of clinically interpretable weighted features. Our findings show that patients diagnosed with schizophrenia may be meaningfully stratified using symptom-based clustering.

Expression of a retinoic acid receptor (RAR)-like protein in the embryonic and adult nervous system of a protostome species.

The vitamin A metabolite, retinoic acid, is an important molecule in nervous system development and regeneration in vertebrates. Retinoic acid signaling in vertebrates is mediated by two classes of nuclear receptors, the retinoid X receptors (RXRs) and the retinoic acid receptors (RARs). Recently, evidence has emerged to suggest that many effects of retinoic acid are conserved between vertebrate and invertebrate nervous systems, even though the RARs were previously thought to be a vertebrate innovation and to not exist in non-chordates. We have cloned a full-length putative RAR from the CNS of the mollusc Lymnaea stagnalis (LymRAR). Immunoreactivity for the RAR protein was found in axons of adult neurons in the central nervous system and in growth cones of regenerating neurons in vitro. A vertebrate RAR antagonist blocked growth cone turning induced by exogenous all-trans retinoic acid, possibly suggesting a role for this receptor in axon guidance. We also provide immunostaining evidence for the presence of RAR protein in the developing, embryonic CNS, where it is also found in axonal processes. Using qPCR, we determined that LymRAR mRNA is detectable in the early veliger stage embryo and that mRNA levels increase significantly during embryonic development. Putative disruption of retinoid signaling in Lymnaea embryos using vertebrate RAR antagonists resulted in abnormal eye and shell development and in some instances completely halted development, resembling the effects of all-trans retinoic acid. This study provides evidence for RAR functioning in a protostome species.

Comparison of hypoglycaemia episodes in people with type-2 diabetes fasting in Ramazan, treated with vildaglipton or sulphonylurea: results of the Pakistani cohort of the VIRTUE study.

OBJECTIVE: To assess the effect of vildagliptin in comparison to sulphonylurea (SU) on hypoglycaemia in Muslim patients with type 2 diabetes mellitus fasting during Ramadan. METHODS: VIRTUE was a multicenter, prospective, observational study, which enrolled 244 patients from Pakistan who were re-analysed. All included patients were treated with vildagliptin (n=121) or SU (n=121) as add-on to metformin or as monotherapy for 16 weeks. The primary outcome of interest was to compare the proportion of patients with ≥1 hypoglycaemic event (HE) during fasting between vildagliptin and SU cohort. Changes in HbA1c and body weight and treatment adherence were also measured. RESULTS: Of the 244 patients enrolled, 120 patients in the vildagliptin cohort (99.2%) and 119 patients in the SU cohort (98.3%) completed the study. Patients experiencing at least one HE were fewer with vildagliptin when compared with SUs (5.8% vs. 14.2%; p<0.033). The reduction in HbA1c was 0.3% with vildagliptin from a baseline of 7.6% and 0.1% with SU from a baseline of 7.4% (between-treatment difference: -0.1% p<0.054). A reduction of 0.3 kg was seen with vildagliptin treatment vs. 0.2 kg weight gain in the SU group. Adverse events were experienced by 15.7% in the vildagliptin cohort and 17.4% in the SU group. CONCLUSION: The treatment with vildagliptin was associated with fewer hypoglycaemic events compared with SUs and was well tolerated with good glycaemic and weight control in patients with T2DM fasting during Ramadan.

Comprehensive watermelon disease recognition dataset.

Plant diseases pose a significant obstacle to global agricultural productivity, impacting crop quality yield and causing substantial economic losses for farmers. Watermelon, a commonly cultivated succulent vine plant, is rich in hydration and essential nutrients. However, it is susceptible to various diseases due to unfavorable environmental conditions and external factors, leading to compromised quality and substantial financial setbacks. Swift identification and management of crop diseases are imperative to minimize losses, enhance yield, reduce costs, and bolster agricultural output. Conventional disease diagnosis methods are often labor-intensive, time-consuming, ineffective, and prone to subjectivity. As a result, there is a critical need to advance research into machine-based models for disease detection in watermelons. This paper presents a large dataset of watermelons that can be used to train a machine vision-based illness detection model. Images of healthy and diseased watermelons from the Mosaic Virus, Anthracnose, and Downy Mildew Disease are included in the dataset's five separate classifications. Images were painstakingly collected on June 25, 2023, in close cooperation with agricultural experts from the highly regarded Regional Horticulture Research Station in Lebukhali, Patuakhali.

Switch from biphasic human insulin 30 to biphasic insulin aspart 30 in Pakistani subjects.

OBJECTIVE: To determine the safety and effectiveness of biphasic insulin aspart 30 (BIAsp 30) in type 2 diabetes subjects switched from biphasic human insulin 30 (BHI 30) in the Pakistani subgroup of the multinational, prospective, non-interventional A1chieve study. METHODS: Subjects who switched therapy from BHI 30 to BIAsp 30 were included in this analysis. Serious adverse drug reactions (SADRs, including major hypoglycaemia) and effectiveness parameters (glycated haemoglobin [HbA1c], fasting plasma glucose [FPG], postprandial plasma glucose [PPPG], systolic blood pressure [SBP]) and body weight were evaluated at the end of 24 weeks. RESULTS: A total of 152 subjects (79 males, 73 females; mean age, 53.4 +/- 10.3 years; BMI, 28.4 +/- 5.8 kg/m2) with an average diabetes duration of 11.2 +/- 4.8 years switched therapy from BHI 30 to BIAsp 30. The mean pre-study BHI 30 dose was 0.66 +/- 0.25 IU/kg and the mean starting BIAsp 30 dose was 0.65 +/- 0.23 U/kg, titrated up to 0.77 +/- 0.22 U/kg after 24 weeks. No SADRs were reported. From baseline to Week 24, overall hypoglycaemia did not change and no major hypoglycaemia was reported at Week 24. HbA1c levels decreased significantly from 9.1 +/- 1.1% at baseline to 7.4 +/- 0.7% (57 +/- 8 mmol/mol) at Week 24 (p < 0.001). Significant improvements in FPG, post-breakfast PPPG and SBP were reported (p < 0.001). CONCLUSION: Switching from BHI 30 to BIAsp 30 was well tolerated and improved glucose control without an increased incidence of hypoglycaemia in this Pakistani cohort.

Entrepreneurial intentions of Gen Z university students and entrepreneurial constraints in Bangladesh.

This research examines a variety of restrictions preventing Bangladeshi youth, particularly Generation Z university students, from becoming involved in entrepreneurship. Moreover, the study examines the influence of Entrepreneurial Attitude (EA), Subjective Entrepreneurial Norms (SEN), Entrepreneurial Perceived Behavioural Control (EPBC), and Entrepreneurial Resilience (ER) on Entrepreneurial Intention (EI) of Bangladeshi Gen Z university students. A systematic literature review methodology following PRISMA procedure was performed to identify the relevant articles. A quantitative method with a positivism philosophy, cross-sectional time horizon and deductive approach was applied to the study. The data of 206 university students from the BBA department of ten universities were collected using convenience sampling and a self-administrated structured questionnaire survey. SPSS 26.0 and Smart PLS 3.0 were used to analyse the data. The output shows a positive and significant association amongst EA, SEN, EPBC, ER, and EI. Various constraints were identified from the literature and ranked based on the respondents' feedback. This research will help entrepreneurs, scholars, policymakers and practitioners to build the entrepreneurial ecosystem and develop young people's understanding of the entrepreneurial decision process and the importance of ER. This paper contributes through empirical investigation to an understanding of the actions that prevent Gen Z students from entrepreneurial activities; decisions are affected by socio-psychological constructions integrating ER with the Theory of Planned behaviour (TPB) model. Triple, Quadruple and Quintuple Helix models are considered supporting theories in this study to shed light on tackling the constraints. To the best knowledge of the researcher, integrating ER with TPB model's constructs is a pioneer scholarly contribution in the context of South-East Asian, specifically Bangladeshi Gen Z students.

Visual Diagnostics of Dental Caries through Deep Learning of Non-Standardised Photographs Using a Hybrid YOLO Ensemble and Transfer Learning Model.

BACKGROUND: Access to oral healthcare is not uniform globally, particularly in rural areas with limited resources, which limits the potential of automated diagnostics and advanced tele-dentistry applications. The use of digital caries detection and progression monitoring through photographic communication, is influenced by multiple variables that are difficult to standardize in such settings. The objective of this study was to develop a novel and cost-effective virtual computer vision AI system to predict dental cavitations from non-standardised photographs with reasonable clinical accuracy. METHODS: A set of 1703 augmented images was obtained from 233 de-identified teeth specimens. Images were acquired using a consumer smartphone, without any standardised apparatus applied. The study utilised state-of-the-art ensemble modeling, test-time augmentation, and transfer learning processes. The "you only look once" algorithm (YOLO) derivatives, v5s, v5m, v5l, and v5x, were independently evaluated, and an ensemble of the best results was augmented, and transfer learned with ResNet50, ResNet101, VGG16, AlexNet, and DenseNet. The outcomes were evaluated using precision, recall, and mean average precision (mAP). RESULTS: The YOLO model ensemble achieved a mean average precision (mAP) of 0.732, an accuracy of 0.789, and a recall of 0.701. When transferred to VGG16, the final model demonstrated a diagnostic accuracy of 86.96%, precision of 0.89, and recall of 0.88. This surpassed all other base methods of object detection from free-hand non-standardised smartphone photographs. CONCLUSION: A virtual computer vision AI system, blending a model ensemble, test-time augmentation, and transferred deep learning processes, was developed to predict dental cavitations from non-standardised photographs with reasonable clinical accuracy. This model can improve access to oral healthcare in rural areas with limited resources, and has the potential to aid in automated diagnostics and advanced tele-dentistry applications.

Psychological benefits of the COVID-19 vaccination: A Bangladeshi comparative study.

Background and Aims: Despite evidence that COVID-19 vaccination can strengthen mental health, there is limited evidence about this in Bangladesh. Thus, this comparative study assessed the prevalence and factors associated with mental health problems between vaccine receivers and nonreceivers. Methods: Using a snowball sampling technique, a web-based cross-sectional study was conducted among a total of 459 participants. The survey questionnaire included sociodemographic information, the Patient Health Questionnaire (PHQ-9), the Generalized Anxiety Disorder (GAD-7), and the Trauma Screening Questionnaire (TSQ-10). Results: The study found that mental health problems were nonsignificantly prevalent in the vaccine nonreceivers than those who received it (i.e., 24.79% vs. 20.60% for depression, 21.20% vs. 16.60% for anxiety, and 15.30% vs. 12.60% for posttraumatic stress disorder). Female gender, chronic condition, smoking status, and alcohol consumption were the risk factors for mental health problems. Conclusion: This study's findings suggest that the COVID-19 vaccination necessarily improves mental health outcomes. However, the study had limitations in terms of its design and sampling technique, and further research is needed to establish a cause-effect relationship between vaccination and mental health problems.

Safety and effectiveness of biphasic insulin aspart 30 (Biasp 30) in people with type 2 diabetes mellitus in the pakistani population: results from the A1chieve study.

OBJECTIVE: To explore the safety and effectiveness of treatment with the insulin analogue, biphasic insulin aspart 30 (BIAsp 30), in people with type 2 diabetes mellitus (T2DM) in a subgroup of a Pakistani population from the A1chieve study. METHODS: A1chieve was a 24-week, international, prospective, multicentre, open label, observational, non-interventional study designed to evaluate the safety and clinical effectiveness of 66,726 people with T2DM who were initiated with basal insulin detemir, fast actinginsulin aspart, and BIAsp 30 (30% soluble insulin aspart, 70% protamine-crystallized insulin aspart). The study was conducted in 28 countries across Asia, Africa, Latin America, and Europe. Here, we report data from a subgroup of 762 people with T2DM from the Pakistani cohort (insulin naïve and insulin users) who were treated withpremix insulin (BIAsp 30) +/- oral antidiabetic drugs (OADs). RESULTS: The decrease in HbAlc at week 24 was statistically significant in the entire cohort, the insulin naïve, and insulin users (1.7 +/- 1.1%, 1.8 +/- 1.3% and 1.7 +/- 0.9%, respectively, p<0.001 for all).There was a statistically significant decrease in the mean fasting plasma glucose (FPG) and postprandial plasma glucose (PPG) from baseline toweek 24 in the entire cohort, in the insulin naïve and in the insulin users with BIAsp 30 treatment (p<0.001 for all).No major hypoglycaemic events were reported during the entire study period. There was a statistically significant decrease in the systolic blood pressure (SBP) in all groups (p<0.001). The improvement in the quality of life score (QoL)was statistically significant in all groups (p<0.001 for all). CONCLUSION: BIAsp 30 treatment appeared to be well tolerated and effective as indicated byimproved glycaemiccontrol and QoL in people with T2DM in the Pakistani population after 24 weeks.

Lifestyle factors associated with the risk of prostate cancer among Pakistani men.

BACKGROUND: Age-adjusted incidence of prostate cancer in Pakistan is 5.3 per 100,000 which is relatively low as compared to other Asian countries, but increasing numbers of cases are being reported. Data on risk factors associated with prostate cancer risk among Pakistani men are sparse. The objective of this study was to identify lifestyle factors associated with the risk of prostate cancer in Pakistani men. METHODS: An unmatched case-control study was conducted in Lahore from February to October 2011. The study enrolled 195 histologically confirmed cases of adenocarcinoma of prostate from Shaukat Khanum Memorial Hospital and Institute of Nuclear Medicine and Oncology Lahore (INMOL) and Lady Reading Hospital. Peshawar, using purposive sampling technique. A total of 390 hospital controls were selected using convenient sampling technique from different teaching hospitals of Lahore after screening with prostate specific antigen levels. A semi-structured interview form was used to collect data through face-to-face interviews. Odds ratio was used as a measure of strength of association and was calculated using unconditional logistic regression. RESULTS: Farmers were found to be at higher odds of prostate cancer (OR 19.76; 95% CI 5.51-70.80; p < 0.001). No significant association was found with marital status, ethnic background, religious affiliation and consanguineous marriages. Level of physical activity was inversely associated with prostate cancer risk (OR 0.05; 95% CI 0.01-0.26; p < 0.001). Positive association was found with increased red meat consumption (OR 11.82; 95% CI 2.88-48.54; p = 0.001) and dairy products intake (11.76; 95% CI 4.23-32.67; p < 0.001). CONCLUSION: Red meat consumption, higher dairy products intake and working as farmers are strongly associated with increased odds of prostate cancer among Pakistani men.

Evaluation of the effectiveness and tolerability of gliclazide modified release 60 mg in patients with type 2 diabetes observing fasting during Ramadan in Pakistan: An analysis from the global DIA-RAMADAN study.

AIM: To assess safety and effectiveness of gliclazide MR 60 mg in people with controlled or suboptimal controlled T2DM treated with breakable gliclazide MR 60 mg formulation. METHOD: This study data has been extracted from an international, observational study conducted in nine Asian and Middle Eastern countries. Total 220 patients with T2DM were recruited from Pakistan. The primary endpoint was the proportion of patients reporting at least 1 symptomatic HE, whereas secondary endpoints were changes in glycosylated haemoglobin (HbA1c) %, fasting plasma glucose (FPG) mg/dL, and body weight (kg) and proportion of patients reporting any HE (confirmed or severe), between inclusion visit (V0) and end of the study visit (V1). RESULTS: During Ramadan, 3.6% (n = 8/220) patients had experienced at least one symptomatic HEs. A significant (p-value < 0.001) reduction was observed in HbA1c: (mean [SD]) (-0.4 [0.9] %), and body weight (-0.7 [4.8] kg). Thirteen adverse events (AEs) unrelated to gliclazide MR were reported during the study pre-Ramadan and post-Ramadan periods. CONCLUSION: This study shows safety and effectiveness profile of gliclazide MR 60 mg by emphasizing on the low risk of HEs, effective glycaemic control and body weight reduction in T2DM patients, who are inclined to fasting during Ramadan.

Automated detection and quantification of Wilms' Tumor 1-positive cells in murine diabetic kidney disease.

In diabetic kidney disease (DKD), podocyte depletion, and the subsequent migration of parietal epithelial cells (PECs) to the tuft, is a precursor to progressive glomerular damage, but the limitations of brightfield microscopy currently preclude direct pathological quantitation of these cells. Here we present an automated approach to podocyte and PEC detection developed using kidney sections from mouse model emulating DKD, stained first for Wilms' Tumor 1 (WT1) (podocyte and PEC marker) by immunofluorescence, then post-stained with periodic acid-Schiff (PAS). A generative adversarial network (GAN)-based pipeline was used to translate these PAS-stained sections into WT1-labeled IF images, enabling in silico label-free podocyte and PEC identification in brightfield images. Our method detected WT1-positive cells with high sensitivity/specificity (0.87/0.92). Additionally, our algorithm performed with a higher Cohen's kappa (0.85) than the average manual identification by three renal pathologists (0.78). We propose that this pipeline will enable accurate detection of WT1-positive cells in research applications.

The Presence and Location of Podocytes in Glomeruli as Affected by Diabetes Mellitus.

The primary purpose of the kidney, specifically the glomerulus, is filtration. Filtration is accomplished through the glomerular filtration barrier, which consists of the fenestrated endothelium, glomerular basement membrane, and specialized epithelial cells called podocytes. In pathologic states, such as Diabetes Mellitus (DM) and diabetic kidney disease (DKD), variable glomerular conditions result in podocyte injury and depletion, followed by progressive glomerular injury and DKD progression. In this work we quantified glomerulus and podocyte structural changes in histopathology image data derived from a murine model of DM. Using a variety of image processing techniques, we studied changes in podocyte morphology and intra-glomerular distribution across healthy, mild DM, and DM glomeruli. Our feature analysis provided feature trends which we believe are reflective of DKD pathology; while glomerular area peaked in mild DM, average podocyte number and distance from the urinary pole continued to decrease and increase, respectively, throughout DM. Ultimately, this study aims to augment the set of quantifiable image biomarkers used for evaluation of DKD progression in digital pathology, as well as underscore the importance of engineering biologically-inspired image features.

Neutrophil Extracellular Traps (NETs): An unexplored territory in renal pathobiology, a pilot computational study.

In the age of modern medicine and artificial intelligence, image analysis and machine learning have revolutionized diagnostic pathology, facilitating the development of computer aided diagnostics (CADs) which circumvent prevalent diagnostic challenges. Although CADs will expedite and improve the precision of clinical workflow, their prognostic potential, when paired with clinical outcome data, remains indeterminate. In high impact renal diseases, such as diabetic nephropathy and lupus nephritis (LN), progression often occurs rapidly and without immediate detection, due to the subtlety of structural changes in transient disease states. In such states, exploration of quantifiable image biomarkers, such as Neutrophil Extracellular Traps (NETs), may reveal alternative progression measures which correlate with clinical data. NETs have been implicated in LN as immunogenic cellular structures, whose occurrence and dysregulation results in excessive tissue damage and lesion manifestation. We propose that renal biopsy NET distribution will function as a discriminate, predictive biomarker in LN, and will supplement existing classification schemes. We have developed a computational pipeline for segmenting NET-like structures in LN biopsies. NET-like structures segmented from our biopsies warrant further study as they appear pathologically distinct, and resemble non-lytic, vital NETs. Examination of corresponding H&E regions predominantly placed NET-like structures in glomeruli, including globally and segmentally sclerosed glomeruli, and tubule lumina. Our work continues to explore NET-like structures in LN biopsies by: 1.) revising detection and analytical methods based on evolving NETs definitions, and 2.) cataloguing NET morphology in order to implement supervised classification of NET-like structures in histopathology images.

High-Mobility Group Box-1 Modulates the Expression of Inflammatory and Angiogenic Signaling Pathways in Diabetic Retina.

PURPOSE: The expression of high-mobility group box-1 (HMGB1) is upregulated in epiretinal membranes and vitreous fluid from patients with proliferative diabetic retinopathy and in the diabetic retina. HMGB1 mediates inflammation, breakdown of the blood-retinal barrier and apoptosis in the diabetic retina. Here, we investigated inflammatory and angiogenic signaling pathways activated by HMGB1 in diabetic retina. METHODS: Human retinal microvascular endothelial cells (HRMEC) and retinas from 1-month diabetic rats and normal rats intravitreally injected with HMGB1 were studied using RT-PCR, Western blot analysis and co-immunoprecipitation. We also studied the effect of the HMGB1 inhibitor glycyrrhizin on diabetes-induced biochemical changes in the retina. RESULTS: Diabetes and intravitreal injection of HMGB1 in normal rats induced significant upregulation of the mRNA levels of the chemokine stromal cell-derived factor-1 (SDF-1/CXCL12) receptor CXCR4 and protein levels of hypoxia-inducible factor-1α, early growth response-1, tyrosine kinase 2 and the CXCL12/CXCR4 chemokine axis. Constant glycyrrhizin intake from onset of diabetes did not affect the metabolic status of the diabetic rats, but it restored these increased mediators to control values. Stimulation of HRMEC with HMGB1 and intraviteral injection of HMGB1 significantly increased the expression of vascular endothelial growth factor (VEGF) and VEGF receptor-2. Co-immunoprecipitation studies showed that diabetes increased the interaction between CXCL12 and CXCR4 and between HMGB1 and receptor for advanced glycation end products (RAGE), but not between HMGB1 and the CXCL12/CXCR4 chemokine axis. CONCLUSIONS: Our findings suggest that HMGB1 activates inflammatory and angiogenic signaling pathways in diabetic retina mediated by RAGE.

Mutual enhancement between high-mobility group box-1 and NADPH oxidase-derived reactive oxygen species mediates diabetes-induced upregulation of retinal apoptotic markers.

The expression of the proinflammatory cytokine high-mobility group box-1 (HMGB1) is upregulated in epiretinal membranes and vitreous fluid from patients with proliferative diabetic retinopathy (PDR) and in the diabetic retina. We hypothesized that a novel mechanism exists where HMGB1 and NADPH oxidase (Nox)-derived reactive oxygen species (ROS) are mutually enhanced in the diabetic retina, which may be a novel mechanism for promoting upregulation of retinal apoptotic markers induced by diabetes. Vitreous samples from 48 PDR and 34 nondiabetic patients, retinas from 1-month diabetic rats and from normal rats intravitreally injected with HMGB1 and human retinal microvascular endothelial cells (HRMEC) stimulated with HMGB1 were studied by enzyme-linked immunosorbent and spectrophotometric assays, Western blot analysis, RT-PCR, and immunofluorescence. We also studied the effect of the HMGB1 inhibitor glycyrrhizin and apocynin on diabetes-induced biochemical changes in the retinas of rats (n = 5-7 in each groups). HMGB1 and the oxidative stress marker protein carbonyl content levels in the vitreous fluid from PDR patients were significantly higher than in controls (p = 0.021; p = 0.005, respectively). There was a significant positive correlation between vitreous fluid levels of HMGB1 and the levels of protein carbonyl content (r = 0.62, p = 0.001). HMGB1 enhanced interleukin-1ß, ROS, Nox2, poly (ADP-ribose) polymerase (PARP)-1, and cleaved caspase-3 production by HRMEC. Diabetes and intravitreal injection of HMGB1 in normal rats induced significant upregulation of ROS, Nox2, PARP-1, and cleaved caspase-3 in the retina. Constant glycyrrhizin and apocynin intake from onset of diabetes did not affect the metabolic status of the diabetic rats, but restored these increased mediators to control values. The results of this study suggest that there is a mutual enhancement between HMGB1 and Nox-derived ROS in the diabetic retina, which may promote diabetes-induced upregulation of retinal apoptotic markers.

Biphasic insulin aspart 30 in insulin-naive people with type 2 diabetes in non-western nations: results from a regional comparative multinational observational study (A(1)chieve).

BACKGROUND: A1chieve(®) (Novo Nordisk A/S, Bagsværd, Denmark) was a prospective, multicenter, noninterventional study in 66,726 people with type 2 diabetes mellitus (T2DM) in 28 countries beginning biphasic insulin aspart 30 (aspart premix), insulin detemir, or insulin aspart in routine clinical care. SUBJECTS AND METHODS: A subgroup of 27,594 insulin-naive people began therapy with aspart premix with or without oral agents. Safety and effectiveness data were taken from clinic records at baseline and after 24 weeks. Seven regional country groupings were prespecified. RESULTS: Mean final insulin dose ranged from 0.68±0.26 U/kg/day (Middle East/Gulf) to 0.38±0.14 U/kg/day (South Asia). The baseline glycated hemoglobin (HbA1c) level varied from 10.5±2.0% (Latin America) to 9.2±1.3% (South Asia), with reductions from -2.9±2.1% (Latin America) to -1.9±1.3% (South Asia). The proportion of people reaching an HbA1c level of <7.0% was highest in China (56%) and lowest in North Africa (22%). Fasting plasma glucose level reductions were from -6.4±5.3 mmol/L (Latin America) to -3.6±2.6 mmol/L (South Asia). Most people began aspart premix twice daily, varying from 91% (North Africa) to 70% (Latin America). Improvement in HbA1c increased with baseline dose frequency (once daily, -1.5±1.4%; twice daily, -2.2±1.6%; three times daily, -2.9±2.2%). CONCLUSIONS: Insulin-naive people with T2DM beginning aspart premix insulin in routine clinical practice in non-western nations had clinically useful improvements in blood glucose control after 24 weeks in all seven regions. Improvements from baseline for glucose control variables were greater than cross-regional differences in those variables at 24 weeks.