Different Stages Of Disease Research Articles

Role of extracellular vesicles in nonalcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD) is a common chronic liver disease that affects approximately one-quarter of the global population and is becoming increasingly prevalent worldwide. The lack of current noninvasive tools and efficient treatment is recognized as a significant barrier to the clinical management of these conditions. Extracellular vesicles (EVs) are nanoscale vesicles released by various cells and deliver bioactive molecules to target cells, thereby mediating various processes, including the development of NAFLD. There is still a long way to actualize the application of EVs in NAFLD diagnosis and treatment. Herein, we summarize the roles of EVs in NAFLD and highlight their prospects for clinical application as a novel noninvasive diagnostic tool as well as a promising therapy for NAFLD, owing to their unique physiochemical characteristics. We summarize the literatures on the mechanisms by which EVs act as mediators of intercellular communication by regulating metabolism, insulin resistance, inflammation, immune response, intestinal microecology, and fibrosis in NAFLD. We also discuss future challenges that must be resolved to improve the therapeutic potential of EVs. The levels and contents of EVs change dynamically at different stages of diseases and this phenomenon may be exploited for establishing sensitive stage-specific markers. EVs also have high application potential as drug delivery systems with low immunogenicity and high biocompatibility and can be easily engineered. Research on the mechanisms and clinical applications of EVs in NAFLD is in its initial phase and the applicability of EVs in NAFLD diagnosis and treatment is expected to grow with technological progress.

Home-based exercise training by using a smartphone app in patients with Parkinson's disease: a feasibility study.

Parkinson's disease (PD) patients experience deterioration in mobility with consequent inactivity and worsened health and social status. Physical activity and physiotherapy can improve motor impairments, but several barriers dishearten PD patients to exercise regularly. Home-based approaches (e.g., via mobile apps) and remote monitoring, could help in facing this issue. This study aimed at testing the feasibility, usability and training effects of a home-based exercise program using a customized version of Parkinson Rehab® application. Twenty PD subjects participated in a two-month minimally supervised home-based training. Daily session consisted in performing PD-specific exercises plus a walking training. We measured: (i) feasibility (training adherence), usability and satisfaction (via an online survey); (ii) safety; (iii) training effects on PD severity, mobility, cognition, and mood. Evaluations were performed at: baseline, after 1-month of training, at the end of training (T2), and at 1-month follow-up (T3). Eighteen out of twenty participants completed the study without important adverse events. Participants' adherence was 91% ± 11.8 for exercise and 105.9% ± 30.6 for walking training. Usability and satisfaction survey scored 70.9 ± 7.7 out of 80. Improvements in PD severity, mobility and cognition were found at T2 and maintained at follow-up. The home-based training was feasible, safe and seems to positively act on PD-related symptoms, mobility, and cognition in patients with mild to moderate stage of PD disease. Additionally, the results suggest that the use of a mobile app might increase the amount of daily physical activity in our study population. Remote monitoring and tailored exercise programs appear to be key elements for promoting exercise. Future studies in a large cohort of PD participants at different stages of disease are needed to confirm these findings.

Возможности диагностического мониторинга диабетической ретинопатии с помощью оптической когерентной томографии-ангиографии

Optical coherence tomography angiography (OCTA) is an advanced noninvasive method of 3D imaging and quantitative analysis of the retinal and choroidal microvasculature. It allows detecting manifestation and progression of diabetic retinopathy, planning treatment and evaluating its results. OCTA expands our understanding of microvascular changes in retinal vascular plexuses in different stages of disease and deepens understanding of its pathogenesis. Keywords: optical coherence tomography angiography; fluorescein angiography; diabetic retinopathy; foveal avascular zone; retinal blood flow

Early chronic suppression of microglial p38α in a model of Alzheimer's disease does not significantly alter amyloid-associated neuropathology.

The p38 alpha mitogen-activated protein kinase (p38α) is linked to both innate and adaptive immune responses and is under investigation as a target for drug development in the context of Alzheimer's disease (AD) and other conditions with neuroinflammatory dysfunction. While preclinical data has shown that p38α inhibition can protect against AD-associated neuropathology, the underlying mechanisms are not fully elucidated. Inhibitors of p38α may provide benefit via modulation of microglial-associated neuroinflammatory responses that contribute to AD pathology. The present study tests this hypothesis by knocking out microglial p38α and assessing early-stage pathological changes. Conditional knockout of microglial p38α was accomplished in 5-month-old C57BL/6J wild-type and amyloidogenic AD model (APPswe/PS1dE9) mice using a tamoxifen-inducible Cre/loxP system under control of the Cx3cr1 promoter. Beginning at 7.5 months of age, animals underwent behavioral assessment on the open field, followed by a later radial arm water maze test and collection of cortical and hippocampal tissues at 11 months. Additional endpoint measures included quantification of proinflammatory cytokines, assessment of amyloid burden and plaque deposition, and characterization of microglia-plaque dynamics. Loss of microglial p38α did not alter behavioral outcomes, proinflammatory cytokine levels, or overall amyloid plaque burden. However, this manipulation did significantly increase hippocampal levels of soluble Aβ42 and reduce colocalization of Iba1 and 6E10 in a subset of microglia in close proximity to plaques. The data presented here suggest that rather than reducing inflammation per se, the net effect of microglial p38α inhibition in the context of early AD-type amyloid pathology is a subtle alteration of microglia-plaque interactions. Encouragingly from a therapeutic standpoint, these data suggest no detrimental effect of even substantial decreases in microglial p38α in this context. Additionally, these results support future investigations of microglial p38α signaling at different stages of disease, as well as its relationship to phagocytic processes in this particular cell-type.

Inflammatory microRNAs in cardiovascular pathology: another brick in the wall.

The regulatory role of microRNAs (miRNAs) is mainly mediated by their effect on protein expression and is recognized in a multitude of pathophysiological processes. In recent decades, accumulating evidence has interest in these factors as modulatory elements of cardiovascular pathophysiology. Furthermore, additional biological processes have been identified as new components of cardiovascular disease etiology. In particular, inflammation is now considered an important cardiovascular risk factor. Thus, in the present review, we will focus on the role of a subset of miRNAs called inflamma-miRs that may regulate inflammatory status in the development of cardiovascular pathology. According to published data, the most representative candidates that play functional roles in thromboinflammation are miR-21, miR-33, miR-34a, miR-146a, miR-155, and miR-223. We will describe the functions of these miRNAs in several cardiovascular pathologies in depth, with specific emphasis on the molecular mechanisms related to atherogenesis. We will also discuss the latest findings on the role of miRNAs as regulators of neutrophil extracellular traps and their impact on cardiovascular diseases. Overall, the data suggest that the use of miRNAs as therapeutic tools or biomarkers may improve the diagnosis or prognosis of adverse cardiovascular events in inflammatory diseases. Thus, targeting or increasing the levels of adequate inflamma-miRs at different stages of disease could help mitigate or avoid the development of cardiovascular morbidities.

Stage Analysis of Breast Cancer Metabolomics: A System Biology Approach.

Breast cancer (BC) is the most common malignancy in women worldwide. Altered miRNA profile can disturb the metabolic homeostatic via regulation of gene expression in BC. In the present study to evaluate which miRNA, regulate metabolic pathways according to their stage, we performed comprehensive analysis of BC expression (mRNA and miRNA) of a set of patients by comparing samples of solid tumor tissue and adjacent tissue. The mRNA and miRNA data of breast cancer were downloaded from the cancer genome database (TCGA) using TCGAbiolinks package. Differentially expressed (mRNAs and miRNAs) was determined by DESeq2 package and predict valid miRNA-mRNA pairs using multiMiR package. All analyses were performed using the R software. Compound-reaction-enzyme-gene network was constructed using the Metscape a plugin for Cytoscape software. Then, core subnetwork computed by CentiScaPe, another plugin for Cytoscape. In Stage I, hsa-miR-592, hsa-miR-449a and hsa-miR-1269a targeted HS3ST4, ACSL1 and USP9Y genes respectively. In stage II, hsa-miR-3662, Hsa-miR-429, and hsa-miR-1269a targeted GYS2, HAS3, ASPA, TRHDE, USP44, GDA, DGAT2, and USP9Y genes. In stage III, hsa-miR-3662 targeted TRHDE, GYS2, DPYS, HAS3, NMNAT2, ASPA genes. In stage IV, hsa-miR-429, has-miR-23c, and hsa-miR-449a targeted genes GDA, DGAT2, PDK4, ALDH1A2, ENPP2, and KL. Those miRNAs and their targets were identified as the discriminative elements for the four stages of breast cancer. The most notable differences between BC and normal tissue in four stages involved multiple pathways and metabolites include: carbohydrate metabolism (e.g., Amylose, N-acetyl-D-glucosamin, beta-D-Glucuronoside, ""g""-CEHC-glucuronide, ""a""-CEHC-glucuronide, Heparan-glucosamine, 5,6-Dihydrouracil, 5,6-Dihydrothymine), branch-chain amino acid metabolism (e.g., N-Acetyl-L-aspartate, N-Formyl-L-aspartate, N`-acetyl-L-asparagine), Retinal metabolism (e.g., Retinal, 9-`cis`-retinal, 13-`cis`-retinal) and (FAD, NAD) as central coenzymes of metabolism. Set of crucial microRNAs and targeted genes plus the related metabolites were introduced for four stages of BC that can be consider for therapeutic and diagnostic purposes in the different stages of disease.

Wearable and digital devices to monitor and treat metabolic diseases.

Cardiometabolic diseases are a major public-health concern owing to their increasing prevalence worldwide. These diseases are characterized by a high degree of interindividual variability with regards to symptoms, severity, complications and treatment responsiveness. Recent technological advances, and the growing availability of wearable and digital devices, are now making it feasible to profile individuals in ever-increasing depth. Such technologies are able to profile multiple health-related outcomes, including molecular, clinical and lifestyle changes. Nowadays, wearable devices allowing for continuous and longitudinal health screening outside the clinic can be used to monitor health and metabolic status from healthy individuals to patients at different stages of disease. Here we present an overview of the wearable and digital devices that are most relevant for cardiometabolic-disease-related readouts, and how the information collected from such devices could help deepen our understanding of metabolic diseases, improve their diagnosis, identify early disease markers and contribute to individualization of treatment and prevention plans.

Three Klebsiella species as potential pathobionts generating endogenous ethanol in a clinical cohort of patients with auto-brewery syndrome: a case control study

Patients with auto-brewery syndrome (ABS) become inebriated after the ingestion of an alcohol-free, high-carbohydrate diet. Our previous work has shown that high-alcohol-producing (HiAlc) Klebsiella pneumoniae can generate excessive endogenous ethanol and cause non-alcoholic fatty liver disease (NAFLD). Therefore, it is reasonable to speculate that such bacteria might play an important role in the pathogenesis of ABS. The characteristics and metabolites of the intestinal flora from a clinical cohort of patients with ABS were analysed during different stages of disease and compared to a group of healthy controls. An invitro culture system of relevant samples was used for screening drug sensitivity and ABS-inducing factors. Rabbit intestinal and murine models were established to verify if the isolated strains could induce ABS invivo. We observed intestinal dysbiosis with decreased abundance of Firmicutes and increased of Proteobacteria in patients with ABS compared with healthy controls. The abundance of the genus Klebsiella in Enterobacteriaceae was strongly associated with fluctuations of patient's blood alcohol concentration. We isolated three species of HiAlc Klebsiella from ABS patients, which were able to induce ABS in mice. Monosaccharide content was identified as a potential food-related inducing factor for alcohol production. Treatments with antibiotics, a complex probiotic preparation and a low-carbohydrate diet not only alleviated ABS, but also erased ABS relapse during the follow-up observation of one of the patients. Excessive endogenous alcohol produced by HiAlc Klebsiella species was an underlying cause of bacterial ABS. Combined prescription of appropriate antibiotics, complex probiotic preparation and a controlled diet could be sufficient for treatment of bacteria-caused ABS. The funders are listed in the acknowledgement.

Co-expression and clinical utility of AR-FL and AR splice variants AR-V3, AR-V7 and AR-V9 in prostate cancer

BackgroundAndrogen receptor (AR) splice variants (AR-Vs) have been discussed as a biomarker in prostate cancer (PC). However, some reports question the predictive property of AR-Vs. From a mechanistic perspective, the connection between AR full length (AR-FL) and AR-Vs is not fully understood. Here, we aimed to investigate the dependence of AR-FL and AR-V expression levels on AR gene activity. Additionally, we intended to comprehensively analyze presence of AR-FL and three clinically relevant AR-Vs (AR-V3, AR-V7 and AR-V9) in different stages of disease, especially with respect to clinical utility in PC patients undergoing AR targeted agent (ARTA) treatment.MethodsAR-FL and AR-V levels were analyzed in PC and non-PC cell lines upon artificial increase of AR pre-mRNA using either drug treatment or AR gene activation. Furthermore, expression of AR-FL and AR-Vs was determined in PC specimen at distinct stages of disease (primary (n = 10) and metastatic tissues (n = 20), liquid biopsy samples (n = 422), mCRPC liquid biopsy samples of n = 96 patients starting novel treatment). Finally, baseline AR-FL and AR-V status was correlated with clinical outcome in a defined cohort of n = 65 mCRPC patients undergoing ARTA treatment.ResultsWe revealed rising levels of AR-FL accompanied with appearance and increase of AR-Vs in dependence of elevated AR pre-mRNA levels. We also noticed increase in AR-FL and AR-V levels throughout disease progression. AR-V expression was always associated with high AR-FL levels without any sample being solely AR-V positive. In patients undergoing ARTA treatment, AR-FL did show prognostic, yet not predictive validity. Additionally, we observed a substantial clinical response to ARTA treatment even in AR-V positive patients. Accordingly, multivariate analysis did not demonstrate independent significance of AR-Vs in neither predictive nor prognostic clinical utility.ConclusionWe demonstrate a correlation between AR-FL and AR-V expression during PC progression; with AR-V expression being a side-effect of elevated AR pre-mRNA levels. Clinically, AR-V positivity relies on high levels of AR-FL, making cells still vulnerable to ARTA treatment, as demonstrated by AR-FL and AR-V positive patients responding to ARTA treatment. Thus, AR-FL and AR-V might be considered as a prognostic, yet not predictive biomarker in mCRPC patients.

Alterations of gut microbes and their correlation with clinical features in middle and end-stages chronic kidney disease.

Gut microecosystem has been shown to play an important role in human health. In recent years, the concept of the gut-kidney axis has been proposed to explain the potential association between gut microbiota and chronic kidney disease (CKD). Here, a cohort of fecal samples collected from patients with CKD (n = 13) were involved. The composition of gut microbial communities and clinical features in CKD and end-stage renal disease (ESRD) were characterized. Our study focused on the changes in gut microbiome and the correlation with clinical features in patients with CKD and ESRD by analyzing high-throughput sequencing results of collected feces. We elucidated the alterations of gut microbiota in CKD patients at different stages of disease and initially identified the gut microbiota associated with CKD progression. We also combined correlation analysis to identify clinical features closely related to the gut microbiome. Our results offered the possibility of using non-invasive gut microbiome in the early diagnosis of course from CKD to ESRD and provide new insights into the association between clinical features and gut microbiota in CKD.

MiSelect R System: the validation of a new detection system of CTCs and their correlation with prognosis in non-metastatic CRC patients

Circulating tumor cells (CTCs) in blood are accepted as a prognostic marker for patients with metastatic colorectal cancer (CRC). However, there is limited data on the use of CTCs as a prognostic marker for non-metastatic patients. In the current study, we used a rare cell automated analysis platform, the MiSelect R System, to enumerate CTCs from blood in non-metastatic CRC patients, and corelated the number of CTCs with the clinical staging and survival. The presence of CTCs in mesenteric vein blood (MVB) samples from 101 CRC patients was significantly associated with T stage. Patients with 1 or more CTCs per 8 mL of MVB exhibited significantly worse disease-free survival (DFS) and cancer-specific survival (CSS) compared to patient without CTCs. The presence of CTCs before surgery is an independent marker for both DFS and CSS. CTC presence after surgical resection is also a prognostic marker. CTCs are a potentially useful prognostic and predictive biomarker in non-metastatic CRC patients that may further stratify patient’s risk status within different stages of disease.

Efficacy of Antibodies Targeting TfR1 in Xenograft Mouse Models of AIDS-Related Non-Hodgkin Lymphoma.

Transferrin receptor 1 (TfR1), also known as CD71, is a transmembrane protein involved in the cellular uptake of iron and the regulation of cell growth. This receptor is expressed at low levels on a variety of normal cells, but is upregulated on cells with a high rate of proliferation, including malignant cells and activated immune cells. Infection with the human immunodeficiency virus (HIV) leads to the chronic activation of B cells, resulting in high expression of TfR1, B-cell dysfunction, and ultimately the development of acquired immunodeficiency syndrome-related B-cell non-Hodgkin lymphoma (AIDS-NHL). Importantly, TfR1 expression is correlated with the stage and prognosis of NHL. Thus, it is a meaningful target for antibody-based NHL therapy. We previously developed a mouse/human chimeric IgG3 specific for TfR1 (ch128.1/IgG3) and showed that this antibody exhibits antitumor activity in an in vivo model of AIDS-NHL using NOD-SCID mice challenged intraperitoneally with 2F7 human Burkitt lymphoma (BL) cells that harbor the Epstein-Barr virus (EBV). We have also developed an IgG1 version of ch128.1 that shows significant antitumor activity in SCID-Beige mouse models of disseminated multiple myeloma, another B-cell malignancy. Here, we aim to explore the utility of ch128.1/IgG1 and its humanized version (hu128.1) in mouse models of AIDS-NHL. To accomplish this goal, we used the 2F7 cell line variant 2F7-BR44, which is more aggressive than the parental cell line and forms metastases in the brain of mice after systemic (intravenous) administration. We also used the human BL cell line JB, which in contrast to 2F7, is EBV-negative, allowing us to study both EBV-infected and non-infected NHL tumors. Treatment with ch128.1/IgG1 or hu128.1 of SCID-Beige mice challenged locally (subcutaneously) with 2F7-BR44 or JB cells results in significant antitumor activity against different stages of disease. Treatment of mice challenged systemically (intravenously) with either 2F7-BR44 or JB cells also showed significant antitumor activity, including long-term survival. Taken together, our results suggest that targeting TfR1 with antibodies, such as ch128.1/IgG1 or hu128.1, has potential as an effective therapy for AIDS-NHL.

Dairy Cows Are Limited in Their Ability to Increase Glucose Availability for Immune Function during Disease.

Shortages of energy and glucose have been hypothesized to play a key role in the development of and responses to production diseases in dairy cows during early lactation. Given the importance of glucose for immune functions, we used a recently established method for the estimation of glucose balance (GB) to evaluate glucose availability during disease phases. A dataset comprising ration analyses as well as individual daily milk yields (MY), dry matter intake (DMI), body weights, and health records of 417 lactations (298 cows) was used to calculate individual daily GB and energy balance (EB). The magnitude and dynamics of MY, DMI, GB, and EB were evaluated in the weeks before, at, and after diagnoses of inflammatory diseases in different stages of early lactation from week in milk 1 to 15. Diagnoses were categorized as mastitis, claw and leg diseases, and other inflammatory diseases. Mixed linear models with a random intercept and slope term for each lactation were used to evaluate the effect of diagnosis on MY, DMI, GB, and EB while accounting for the background effects of week in milk, parity, season, and year. When unaffected by disease, in general, the GB of cows was close to zero in the first weeks of lactation and increased as lactation progressed. Weekly means of EB were negative throughout all lactation stages investigated. Disease decreased both the input of glucose precursors due to a reduced DMI as well as the output of glucose via milk due to a reduced MY. On average, the decrease in DMI was -1.5 (-1.9 to -1.1) kg and was proportionally higher than the decrease in MY, which averaged -1.0 (-1.4 to -0.6) kg. Mastitis reduced yield less than claw and leg disease or other diseases. On average, GB and EB were reduced by -3.8 (-5.6 to -2.1) mol C and -7.5 (-10.2 to -4.9) MJ in the week of diagnosis. This indicates the need to investigate strategies to increase the availability of glucogenic carbon for immune function during disease in dairy cows.

Data Analysis of Impaired Renal and Cardiac Function Using a Combination of Standard Classifiers

We examine the significance of the predictive potential of EPI cystatin C (EPI CysC) in combination with NTproBNP, sodium, and potassium in the evaluation of renal function in patients with cardiorenal syndrome using standard mathematical classification models from the domain of artificial intelligence. The criterion for the inclusion of subjects with combined impairment of heart and kidney function in the study was the presence of newly discovered or previously diagnosed clinically manifest cardiovascular disease and acute or chronic kidney disease in different stages of evolution. In this paper, five standard classifiers from the field of machine learning were used for the analysis of the obtained data: ensemble of neural networks (MLP), ensemble of k-nearest neighbors (k-NN) and naive Bayes classifier, decision tree, and a classifier based on logistic regression. The results showed that in MLP, k-NN, and naive Bayes, EPI CysC had the highest predictive potential. Thus, our approach with utility classifiers recognizes the essence of the disorder in patients with cardiorenal syndrome and facilitates the planning of further treatment.

Common mouse models of tauopathy reflect early but not late human disease

BackgroundMouse models that overexpress human mutant Tau (P301S and P301L) are commonly used in preclinical studies of Alzheimer’s Disease (AD) and while several drugs showed therapeutic effects in these mice, they were ineffective in humans. This leads to the question to which extent the murine models reflect human Tau pathology on the molecular level.MethodsWe isolated insoluble, aggregated Tau species from two common AD mouse models during different stages of disease and characterized the modification landscape of the aggregated Tau using targeted and untargeted mass spectrometry-based proteomics. The results were compared to human AD and to human patients that suffered from early onset dementia and that carry the P301L Tau mutation.ResultsBoth mouse models accumulate insoluble Tau species during disease. The Tau aggregation is driven by progressive phosphorylation within the proline rich domain and the C-terminus of the protein. This is reflective of early disease stages of human AD and of the pathology of dementia patients carrying the P301L Tau mutation. However, Tau ubiquitination and acetylation, which are important to late-stage human AD are not represented in the mouse models.ConclusionAD mouse models that overexpress human Tau using risk mutations are a suitable tool for testing drug candidates that aim to intervene in the early formation of insoluble Tau species promoted by increased phosphorylation of Tau.

3D-printed immunosensor for the diagnosis of Parkinson’s disease

3D printing technology is a strategic tool for the development of electrochemical sensors and biosensors since it is possible to obtain versatile devices quickly and at a low cost. In this work, an arrangement of 3D-printed electrodes (working, pseudo-reference, and auxiliary) was applied for the detection of PARK7/DJ-1 protein in blood serum and cerebrospinal fluid samples. The immunosensor surface was previously chemically and electrochemically activated to promote the increase of the active sites and the conductivity, allowing the covalent immobilization of the biological species (antibodies) and improving its electrochemical performance. The detection was carried out by impedimetric (5.0 −200 µg L−1), and voltammetric measurements (5.0 −500 µg L−1), showing limits of detection of 1.01 and 3.46 µg L−1. The 3D-printed immunosensor also achieved good repeatability and reproducibility from normal to abnormal levels of PARK7/DJ-1 protein, aiming for the diagnosis of Parkinson’s disease in different stages of the disease.

Helicobacter pylori Chronic Infection Selects for Effective Colonizers of Metaplastic Glands.

Chronic gastric infection with Helicobacter pylori can lead to progressive tissue changes that culminate in cancer, but how H. pylori adapts to the changing tissue environment during disease development is not fully understood. In a transgenic mouse gastric metaplasia model, we found that strains from unrelated individuals differed in their ability to infect the stomach, to colonize metaplastic glands, and to alter the expression of the metaplasia-associated protein TFF3. H. pylori isolates from different stages of disease from a single individual had differential ability to colonize healthy and metaplastic gastric glands. Exposure to the metaplastic environment selected for high gastric colonization by one of these strains. Complete genome sequencing revealed a unique alteration in the frequency of a variant allele of the putative adhesin sabB, arising from a recombination event with the related sialic acid binding adhesin (SabA) gene. Mutation of sabB in multiple H. pylori strain backgrounds strongly reduced adherence to both normal and metaplastic gastric tissue, and highly attenuated stomach colonization in mice. Thus, the changing gastric environment during disease development promotes bacterial adhesin gene variation associated with enhanced gastric colonization. IMPORTANCE Chronic infection with Helicobacter pylori is the primary risk factor for developing stomach cancer. As disease progresses H. pylori must adapt to a changing host tissue environment that includes induction of new cell fates in the cells that line the stomach. We tested representative H. pylori isolates collected from the same patient during early and later stages of disease in a mouse model where we can rapidly induce disease-associated tissue changes. Only the later-stage H. pylori strains could robustly colonize the diseased stomach environment. We also found that the ability to colonize the diseased stomach was associated with genetic variation in a putative cell surface adhesin gene called sabB. Additional experiments revealed that SabB promotes binding to stomach tissue and is critical for stomach colonization by the late-stage strains. Thus, H. pylori diversifies its genome during disease progression and these genomic changes highlight critical factors for bacterial persistence.

AYURVEDIC TREATMENT OF DIABETIC RETINOPATHY-A SINGLE CASE STUDY

Diabetic Retinopathy refers to a complication of diabetes that affects the eyes. That is nothing but damage to blood vessels in the tissue of retina. With increase in the life expectancy of diabetics, the incidence of Diabetic Retinopathy has increased. (1) Diabetic Retinopathy is leading cause of blindness. There are many treatment modalities mentioned in modern medicine but it have certain limitation. In Ayurveda, Diabetic Retinopathy can be compared with Madhumehajanya Timir. All the three doshas along with Rakta dosha and sapta dhatu with four internal Drishti patalas of eyes are affected in Madhumehajanya Timir in different stages of disease. In this current case of Madhumehajanya Timir, Ayurvedic treatment was done. In present case study a female patient of 58 years old visited the OPD of Shalakyatantra with complaining of defective distant and near vision since 6 months. Based upon the history and clinical features, Patient is K/C/O diabetes mellitus (type 2) since 8 years and is on regular medication. Patient is treated with Vasant Kusumakar Rasa 1OD for 1 month ,Tab BGR-34 2BD for 6 month, Tab Saptamrit Lauh 500 mg 1BD for 1 month and Triphala Ghrita 10 ml HS for 3 months duration. At the end of 6 months patient got significant relief.

IMPAIRED BALANCE OF CLOTTING FACTORS IN LIVER CIRRHOSIS

Liver cirrhosis (LC) has been accepted as prototype of a disease with acquired prohemorrhagic diathesis. However, as the synthesis of all coagulant factors is affected, a rebalanced but fragile hemostasis is maintained. With increasing disease severity, a disproportion between levels of certain coagulants occurs that can lead to prothrombotic tendency. We aimed to evaluate the levels of factor VIII (FVIII), protein C (PC) and their ratio (FVIII/PC) as the main determinants of thrombin generation in patients with LC at different stages of disease. Fifty patients with LC were divided in three groups according to LC severity using the Child-Turcotte-Pugh Score (CTP-A, CTP-B, CTP-C). The levels of FVIII and protein C were measured in sodium citrate plasma on Siemens, BCS XP Blood Coagulometer. The levels of FVIII, PC and FVIII/PC were compared between the groups and a correlation of their values to MELD score was performed. Plasma levels of FVIII increased with severity of the disease, with concurrent statistically significant decrease of plasma PC levels (p=0.0008). This was accompanied with statistically significant increase of the FVIII/PC ratio (p=0.0004) indicating hypercoagulable state in advanced stage of the disease. A significant correlation to MELD score was identified for PC in group CTP-B and CTP-C and for FVIII/PC ratio in group CTP-C. As liver cirrhosis severity increases, a disproportion in plasma levels of the most powerful determinants of thrombin generation occurs. This could be the explanation for the observed increased risk of venous thromboembolism in patients with liver cirrhosis.

Sensitive SARS-CoV-2 detection, air travel Covid-19 testing, variant determination and fast direct PCR detection, using ddPCR and RT-qPCR methods.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) monitoring in air traffic is important in the prevention of the virus spreading from abroad. The gold standard for SARS-CoV-2 detection is RT-qPCR; however, for early and low viral load detection, a much more sensitive method, such as droplet digital PCR (ddPCR), is required. Our first step was to developed both, ddPCR and RT-qPCR methods, for sensitive SARS-CoV-2 detection. Analysis of ten swab/saliva samples of five Covid-19 patients in different stages of disease showed positivity in 6/10 samples with RT-qPCR and 9/10 with ddPCR. We also used our RT-qPCR method for SARS-CoV-2 detection without the need of RNA extraction, obtaining results in 90-120 minutes. We analyzed 116 self-collected saliva samples from passengers and airport staff arriving from abroad. All samples were negative by RT-qPCR, while 1 was positive, using ddPCR. Lastly, we developed ddPCR assays for SARS-CoV-2 variants identification (alpha, beta, gamma, delta/kappa) that are more economically advantageous when compared to NGS. Our findings demonstrated that saliva samples can be stored at ambient temperature, as we did not observe any significant difference between a fresh sample and the same sample after 24 hours (p = 0.23), hence, saliva collection is the optimal route for sampling airplane passengers. Our results also showed that droplet digital PCR is a more suitable method for detecting virus from saliva, compared to RT-qPCR. Keywords: COVID-19; RT-PCR; ddPCR; SARS-CoV-2; nasopharyngeal swab; saliva.