Extracellular Vesicles Contribute to Viral-Induced Hepatocellular Carcinoma: Understanding Their Involvement in Viral Hepatitis and Their Potential as Biomarkers for Early Hepatocellular Carcinoma Detection.

The high global prevalence of hepatitis B and hepatitis C and the poor prognosis of hepatitis B and hepatitis C-associated hepatocellular carcinoma (HCC), necessitates the early diagnosis and treatment of the disease. Recent studies show that cell-to-cell communication via extracellular vesicles (EVs) is involved in the HCC progression. The objective of the following study was to explore the role of EVs in the progression of viral-induced HCC and investigate their potential for the early diagnosis of cancer. First, the mRNA derived from EVs of HCC patients was compared to the mRNA derived from EVs from the healthy controls. Expression analysis of ANGPTL3, SH3BGRL3, and IFITM3 genes from the EVs was done. Afterward, to confirm whether hepatocytes can uptake EVs, HuH7 cells were exposed to EVs, and the expression analysis of downstream target genes (AKT, TNF-α, and MMP-9) in Huh7 cells was done. Transcriptional analysis showed that in the EVs from HCC patients, the expression levels of ANGPTL3, SH3BGRL3, and IFITM3 were significantly increased by 2.62-, 4.3-, and 9.03-folds, respectively. The downstream targets, AKT, TNF-α, and MMP-9, also showed a considerable change of 4.1-, 1.46-, and 5.05-folds, respectively, in Huh7 cells exposed to HCC EVs. In conclusion, the following study corroborates the role of EVs in HCC progression. Furthermore, the significant alteration in mRNA levels of the selected genes demonstrates their potential to be used as possible biomarkers for the early diagnosis of HCC.

The potential of IFN-λ, IL-32γ, IL-6, and IL-22 as safeguards against human viruses: a systematic review and a meta-analysis.

Many studies have investigated the antiviral activity of cytokines, including interleukin-6 (IL-6), interleukin-22 (IL-22), interleukin-32 gamma (IL-32γ), and interferon-lambda (IFN-λ) in diverse populations. This study aims to evaluate the role of these cytokines in inhibition of various human and animal viruses when administered exogenously. A comprehensive meta-analysis and systematic review were conducted on all the relevant studies from three databases. Standard mean differences (SMDs) of overall viral inhibition were used to generate the difference in the antiviral efficacy of these cytokines between control and experimental groups. A total of 4,618 abstracts for IL-6, 3,517 abstracts for IL-22, 2,160 abstracts for IL-32γ, and 1,026 abstracts for IFN-λ were identified, and 7, 4, 8, and 35 studies were included, respectively, for each cytokine. IFN-λ (SMD = 0.9540; 95% CI: 0.69-0.22) and IL-32γ (SMD = 0.459; 95% CI: 0.02-0.90) showed the highest influence followed by IL-6 (SMD = 0.456; CI: -0.04-0.95) and IL-22 (SMD = 0.244; 95% CI: -0.33-0.81). None of the cytokines represented heterogeneity (tau² > 0), but only IFN-λ indicated the funnel plot asymmetry (p = 0.0097). Results also indicated that IFN-λ and IL-32γ are more potent antivirals than IL-6 and IL-22. The collective findings of this study emphasize that exogenously administered pro-inflammatory cytokines, specifically IFN-λ and IL-32, exhibit a significant antiviral activity, thereby underscoring them as potent antiviral agents. Nonetheless, additional research is required to ascertain their clinical utility and potential for integration into combinatorial therapeutic regimens against viral infections.

Antioxidants and Calcium Modulators Preclude in Vitro Hepatitis B Virus-Induced Mitochondrial Damage.

BACKGROUND/AIMS: Hepatitis B virus induces mitochondrial damage via the production of reactive oxygen species and concomitant with deregulation of calcium homeostasis. The current study evaluates the potential of antioxidant and calcium modulators for inhibition of hepatitis B virus-induced mitochondrial damage using in vitro cell culture models. MATERIALS AND METHODS: Hepatitis B virus-induced mitochondrial fragmentation was observed by immunofluorescence confocal micros- copy in hepatitis B virus-infected cell lines (HepG2 and HepAD38). Differential protein expression of mitochondrial fragmentation mark- ers, dynamin-related protein 1 and phospho-dynamin-related protein 1, were evaluated both pre- and posttreatment with antioxidant N-acetyl-l-cysteine and calcium modulators like 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakisacetoxymethyl ester, ethylene-bis (oxyethylenenitrilo) tetraacetic acid glycol ether diamine tetraacetic acid-acetoxymethyl ester, and ruthenium amine complex by western blot analysis. RESULTS: A slight reduction in mitochondrial fragmentation in both cell lines was observed post-antioxidant treatment with a partial prevention observed with calcium modulators. The expression of phospho-dynamin-related protein 1 was significantly upregulated (P = .0007, P = .003) in both hepatitis B virus-infected cell lines compared to uninfected cells. In line with these observations, the expres- sion of dynamin-related protein 1 and phospho-dynamin-related protein 1 was found to be significantly downregulated with N-acetyl- l-cysteine treatment in both cell lines (P = .003, P = .002), respectively. A nonsignificant trend was observed in the case of calcium modulators treatment. CONCLUSIONS: Current study indicates that the mitochondrial fragmentation induced by hepatitis B virus infection can be reduced after antioxidant treatment pointing toward exploring better drug targets for the prevention of hepatitis B virus-induced mitochondrial frag- mentation and associated liver damage.

Interleukin-22 promotes the proliferation and migration of hepatocellular carcinoma cells via the phosphoinositide 3-kinase (PI3K/AKT) signaling pathway.

BACKGROUND: Interleukin-22 (IL-22) is a pro-inflammatory cytokine released during the immune response in chronic liver injury. Although IL-22 mediates tissue regeneration, its uncontrolled production may generate a carcinogenic environment resulting in hepatocellular carcinoma (HCC). This study aims to identify the effect of IL-22 on anti-apoptotic and metastatic genes and the molecular pathways responsible for IL-22-mediated hepatic carcinogenesis. METHODS AND RESULTS: Three cancerous liver lines, HepG2, SNU-387, Huh7, and one normal liver line, THLE2, were treated with IL-22. RT-qPCR analysis was conducted to study the role of IL-22 in altering the expression levels of anti-apoptotic genes, MCL-1 and BCL-2, and metastatic genes, MMP-7 and MMP-9. A significant increase in expression levels of these genes was observed after IL-22 treatment. Furthermore, to explore the major pathways involved in IL-22-mediated upregulation of anti-apoptotic and metastatic genes, cells were treated with inhibitors of JAK/STAT and PI3K/AKT pathways along with IL-22. Resultantly, a significant decrease in expression levels of target genes was observed, indicating the involvement of JAK/STAT and PI3K/AKT signaling cascades in IL-22-mediated oncogenesis. Finally, Cell Scratch assay was performed to check the effect of IL-22 and inhibitors of JAK/STAT and PI3K/AKT on the metastatic potential of liver cells. While migration was observed in Huh7 and THLE2 cells treated with IL-22, no migration was observed in cells treated with IL-22 along with JAK/STAT and PI3K/AKT inhibitors. Results indicate that IL-22 encourages metastasis in HCC cells via the JAK/STAT and PI3K/AKT pathways. CONCLUSION: Results showed that IL-22 upregulates anti-apoptotic and metastatic genes in HCC through JAK/STAT and PI3K/AKT signaling pathways.

Navigating the brain: the role of exosomal shuttles in precision therapeutics.

Brain diseases have become one of the leading roots of mortality and disability worldwide, contributing a significant part of the disease burden on healthcare systems. The blood-brain barrier (BBB) is a primary physical and biological obstacle that allows only small molecules to pass through it. Its selective permeability is a significant challenge in delivering therapeutics into the brain for treating brain dysfunction. It is estimated that only 2% of the new central nervous system (CNS) therapeutic compounds can cross the BBB and achieve their therapeutic targets. Scientists are exploring various approaches to develop effective cargo delivery vehicles to promote better therapeutics targeting the brain with minimal off-target side effects. Despite different synthetic carriers, one of the natural brain cargo delivery systems, "exosomes," are now employed to transport drugs through the BBB. Exosomes are naturally occurring small extracellular vesicles (EVs) with unique advantages as a therapeutic delivery system for treating brain disorders. They have beneficial innate aspects of biocompatibility, higher stability, ability to cross BBB, low cytotoxicity, low immunogenicity, homing potential, targeted delivery, and reducing off-site target effects. In this review, we will discuss the limitations of synthetic carriers and the utilization of naturally occurring exosomes as brain-targeted cargo delivery vehicles and highlight the methods for modifying exosome surfaces and drug loading into exosomes. We will also enlist neurodegenerative disorders targeted with genetically modified exosomes for their treatment.

Development of novel antiviral nanofinishes for bioactive textiles.

Virus-caused public health outbreaks represent a serious threat to humans all over the world. The rampant new 2019 coronavirus (SARS-CoV-2) has wreaked havoc on China and the rest of the world since December 2019. Now focus is on effective reduction of corona and other viral and bacterial infections in hospitals, public and private sectors, households, schools, etc. Metal and metal oxide nanoparticles, carbon nanotubes, heterostructures, patterned surfaces, and graphene-based materials have shown up to 99.9998% efficacy against bacteria, mold, and viruses. The stability, long shelf life, and robustness of inorganic nanoparticles make them desirable for antimicrobial nanofinishes. These inorganic antimicrobial agents are more stable than organic antibacterial compounds at high temperature and pressure. The high specific surface area-to-volume ratios and unique physicochemical characteristics of nanoparticles are largely responsible for their antibacterial actions. But their immobilization is a huge challenge. To address this issue, NPs were modified with (glycidoxypropyl) trimethoxysilane (GPTS) and applied on cotton fabric. The silane part of GPTS reacted with the NPs under acidic conditions while epoxy reacted with cotton under alkaline conditions. Treated cotton fabric showed good antiviral and antibacterial activity even after severe industrial washing.

Emergence and Persistence of Resistance-Associated Substitutions in HCV GT3 Patients Failing Direct-Acting Antivirals.

Background: The hepatitis C virus has a high mutation rate, which results in the emergence of resistance-associated substitutions (RASs). Despite direct-acting antivirals (DAAs) efforts to treat chronically infected HCV genotype 3 (GT3) patients, there are concerns about the emergence and persistence of RASs in DAA failures. The objective of this study was to determine the prevalence of clinically relevant RASs in HCV NS5A and NS5B regions before and after treatment to better understand the role of RASs in treatment failures. Methods: Viral RNA was extracted before and after treatment from serum samples. NS5A and NS5B regions of HCV were amplified by nested PCR, followed by Sanger sequencing. The nucleotide sequences were aligned against HCV GT3 reference sequences, and amino acid substitutions were analyzed using the geno2pheno [hcv] webserver. Results: A total of 76 patients failing DAA therapy were stratified from the cohort of 1388. RASs were detected at the baseline in 15/76 patients and at relapse in 20/76 patients with cirrhosis and previously treated with interferons. The most prevalent NS5A RAS was Y93H found in all treatment-failing patients (14/54 in DCV vs. 6/22 in VEL), followed by A62S/T and A30K. No RASs were identified in NS5B. RASs that were present at the baseline persisted through the 24-week follow-up period and were enriched with emerging RASs during the treatment. The presence of RASs may be one of the causes of treatment failures in 26.3% of patients. Amino acid substitutions were present at the baseline in most of the patients with RASs against NS5A inhibitors. Patients with the baseline Y93H and/or A30K relapse more frequently than patients harboring A62S/T. Conclusion: Treatment-failing patients harbored NS5A RASs, and the most frequent were A30K (5/20), A62S/T (20/20), and Y93H (20/20). Direct resistance testing is recommended for optimizing re-treatment strategies in treatment-failing patients.

P2X4 receptors mediate induction of antioxidants, fibrogenic cytokines and ECM transcripts; in presence of replicating HCV in in vitro setting: An insight into role of P2X4 in fibrosis.

BACKGROUND & AIMS: Major HCV infections lead to chronic hepatitis, which results in progressive liver disease including fibrosis, cirrhosis and eventually hepatocellular carcinoma (HCC). P2X4 and P2X7 are most widely distributed receptors on hepatocytes. METHODS: Full length P2X4 (1.7kb) (Rattus norvegicus) was sub cloned in mammalian expression vector pcDNA3.1+. Two stable cell lines 293T/P2X4 (experimental) and 293T/ NV or null vector (control) were established. Both cell lines were inoculated with high viral titers human HCV sera and control human sera. Successfully infected cells harvested on day 5 and day 9 of post infection were used for further studies. RESULTS: The results revealed a significant increase in gene expression of P2X4 on day 5 and day 9 Post -infection in cells infected with HCV sera compared with cells inoculated with control sera. Quantitative real time PCR analysis revealed that HO-1 was significantly upregulated in presence of P2X4 in HCV infected cells (P2X4/HCV) when compared with control NV/HCV cells. A significant decrease was observed in expression of Cu/ZnSOD in presence of P2X4 in HCV infected cells compared to control NV/HCV cells. However, expression of both antioxidants was observed unaltered in cells harvested on day 9 post infection. Gene expression of angiotensin II significantly increased in HCV infected cells in presence of P2X4 on day 5 and day 9 of post infection when compared with control NV/HCV cells. A significant increase in gene expression of TNF-α and TGF-ß was observed in HCV infected cells in presence of P2X4 on day 9 post infection in comparison with control (NV/HCV cells). However, gene expression of adipokine leptin was not affected in both experimental (P2X4/HCV) and control (NV/HCV) groups on day 5 and day 9 of post infection. Extracellular matrix proteins, laminin and elastin genes expression also significantly increased in presence of P2X4 (HCV/P2X4) on day 9 of post-infection compared to control group NV/HCV cells. CONCLUSION: In conclusion, these findings constitute the evidence that P2X4 receptors in the presence of HCV play a significant role in the regulation of key antioxidant enzymes (HO-1, Cu/ZnSOD), in the induction of proinflammatory. cytokine (TNF-α), profibrotic cytokine (TGF-ß) vasoactive cytokine (angiotensin II). P2X4 also increases the expression of extracellular matrix proteins (laminin and elastin) in the presence of HCV.

WITHDRAWN: Original Antigenic Sin in COVID-19: Hoskins Effect and Vaccine

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The global emergence of Chikungunya infection: An integrated view.

Chikungunya virus (CHIKV) is one of the emerging viruses around the globe. It belongs to the family Togaviridae and genus Alphavirus and is an arthropod borne virus that transmits by the bite of an infected mosquito, mainly through Aedes aegypti and Aedes albopcitus. It is a spherical, enveloped virus with positive single stranded RNA genome. It was first discovered during 1952-53 in Tanganyika, after which outbreaks were documented in many regions of the world. CHIKV has two transmission cycles; an enzootic sylvatic cycle and an urban cycle. CHIKV genome contains 11,900 nucleotides and two open reading frames and shows great sequence variability. Molecular mechanisms of virus host-cell interactions and the pathogenesis of disease are not fully understood. The disease involves three phases; acute, post-acute and chronic with symptoms including high-grade fever, arthralgia, macupapular rashes and headache. There is no licensed vaccine or specific treatment for CHIKV infection. This lack of specific interventions combined with difficulties in making a precise diagnosis together make the disease difficult to manage. In this review we aim to present the current knowledge of global epidemiology, transmission, structure, various aspects of diagnosis as well as highlight potential antiviral drugs and vaccines against CHIKV.

Unilateral absence of external oblique musculo-aponeurotic complex during routine inguinal hernia repair.

INTRODUCTION: The isolated absence of the external oblique musculo-aponeurotic complex in adults is an extremely rare anomaly. This complex is an important contributor to the strength of the inguinal canal. The present case report describes the unilateral absence of the external oblique muscle in a patient. CASE PRESENTATION: A 40-year-old male patient presented with a history of intermittent lower abdominal pain for 5 years which had increased over the past 6 months. Abdominal examination revealed unilateral reducible, incomplete, left sided direct inguinal hernia. CLINICAL DISCUSSION: Elective unilateral Lichtenstein's mesh hernioplasty was planned for the patient. Intraoperatively, there was no evidence of the external oblique aponeurosis and the spermatic cord was noted deep to the thickened membranous fascial layer. The inguinal ligament was thin and atrophic and was attached to the pubic tubercle medially and anterior superior iliac spine laterally. There was no evidence of any superior aponeurotic connection to the inguinal ligament. A postoperative ultrasound examination of the abdomen confirmed the unilateral absence of the external oblique musculo-aponeurotic complex. CONCLUSION: The possibility of such an anomaly should be considered in patients without other risk factors for hernia.

Biochar and slow-releasing nitrogen fertilizers improved growth, nitrogen use, yield, and fiber quality of cotton under arid climatic conditions.

The efficiency of nitrogenous fertilizers in South Asia is on a declining trajectory due to increased losses. Biochar (BC) and slow-releasing nitrogen fertilizers (SRNF) have been found to improve nitrogen use efficiency (NUE) in certain cases. However, field-scale studies to explore the potential of BC and SRNF in south Asian arid climate are lacking. Here we conducted a field experiment in the arid environment to demonstrate the response of BC and SRNF on cotton growth and yield quality. The treatments were comprised of two factors, (A) nitrogen sources, (i) simple urea, (ii)neem-coated urea, (iii)sulfur-coated urea, (iv) bacterial coated urea, and cotton stalks biochar impregnated with simple urea, and (B) nitrogen application rates, N1=160 kg ha-1, N2 = 120 kg ha-1, and N3 = 80 kg ha-1. Different SRNF differentially affected cotton growth, morphological and physiological attributes, and seed cotton yield (SCY). The bacterial coated urea at the highest rate of N application (160 kg ha-1) resulted in a higher net leaf photosynthetic rate (32.8 µmol m-2 s-1), leaf transpiration rate (8.10 mmol s-1), and stomatal conductance (0.502 mol m-2 s-1), while leaf area index (LAI), crop growth rate (CGR), and seed cotton yield (4513 kg ha-1) were increased by bacterial coated urea at 120 kg ha-1 than simple urea. However, low rate N application (80 kg ha-1) of bacterial coated urea showed higher nitrogen use efficiency (39.6 kg SCY kg-1 N). The fiber quality (fiber length, fiber strength, ginning outturn, fiber index, and seed index) was also increased with the high N application rates than N2 and N3 application. To summarize, the bacterial coated urea with recommended N (160 kg ha-1) and 75% of recommended N application (120 kg ha-1) may be recommended for farmers in the arid climatic conditions of Punjab to enhance the seed cotton yield, thereby reducing nitrogen losses.

Platelets' RNA as biomarker trove for differentiation of early-stage hepatocellular carcinoma from underlying cirrhotic nodules.

BACKGROUND & AIMS: Among the multiplicity of factors involved in rising incidence of hepatocellular carcinoma (HCC)-the second deadliest cancer, late diagnosis of early-stage HCC nodules originating from late-stage cirrhotic nodules is the most crucial. In recent years, Tumor-educated platelets (TEPs) have emerged as a strong multimodal tool to be used in liquid-biopsy of cancers because of changes in their mRNA content. This study assessed the reliability of selected mRNA repertoire of platelets as biomarkers to differentiate early HCC from late-stage cirrhotic nodules. METHODS: Quantitative real time PCR (qRT-PCR) was used to evaluate expression levels of selected platelets-specific mRNA between HCC patients compared to cirrhosis patients. ROC curve analysis assessed the sensitivity and specificity of the biomarkers. RESULTS: RhoA, CTNNB1 and SPINK1 showed a significant 3.3-, 3.2- and 3.18-folds upregulation, respectively, in HCC patients compared to cirrhosis patients while IFITM3 and SERPIND1 presented a 2.24-fold change. Strikingly, CD41+ platelets also demonstrated a marked difference of expression in HCC and cirrhosis groups. CONCLUSIONS: Our study reports liquid biopsy-based platelets mRNA signature for early diagnosis of HCC from underlying cirrhotic nodules. Moreover, differential expression of CD41+ platelets in two groups provides new insights into a probable link between CD41 expression on platelets with the progression of cirrhosis to HCC.

A comparative analysis of interferons and direct-acting antivirals on the expression of genes involved in hepatitis C pathogenesis.

The discovery of direct-acting antivirals (DAAs) has revolutionized the treatment of hepatitis C worldwide. In contrast, pegylated interferon-alpha (PEG IFN-α), the older regimen, had limited success. However, the effect of DAAs on the expression of immunomodulatory genes involved in liver pathologies remains ambiguous. The objective of this study was to explore and contrast the effects of DAAs and PEG IFN-α on the expression of selected immunomodulatory genes. Fifty individuals were enrolled in the study and they were divided into five categories; healthy individuals, treatment-naive, DAAs-responders, DAAs-nonresponders, and interferon-relapsers. The effect of the therapies on the expression of transforming growth factor-beta (TGF-ß), tumor necrosis factor-alpha (TNF-α), suppressor of cytokine signaling 3 (SOCS-3), copper/zinc superoxide dismutase (Cu/Zn SOD), interleukin 10 (IL-10), and collagen type 1 was analyzed. Expression analysis of the selected genes was done through real time polymerase chain reaction. A significantly increased expression of TGF-ß was observed in the patients who received DAAs or PEG IFN-α, which suggests that patients receiving anti-HCV therapies are prone to developing fibrosis. Moreover, DAAs-nonresponders had higher expression of TNF-α, SOCS-3, and IL-10. The elevated expression of TNF-α and SOCS-3 insinuates that DAAs-nonresponders may develop insulin resistance and steatosis in the future. Finally, in addition to TGF-ß, high expression of collagen was found in interferon relapsers, which suggests that these patients are the most susceptible to developing cirrhosis.

A Double Edged Sword Role of Interleukin-22 in Wound Healing and Tissue Regeneration.

Wound healing and tissue regeneration is an intricate biological process that involves repair of cellular damage and maintenance of tissue integrity. Cascades involved in wound healing and tissue regeneration highly overlap with cancer causing pathways. Usually, subsequent tissue damage events include release of a number of cytokines to accomplish post-trauma restoration. IL-22 is one of the cytokines that are immediately produced to initiate immune response against several tissue impairments. IL-22 is a fundamental mediator in inflammation, mucous production, protective role against pathogens, wound healing, and tissue regeneration. However, accumulating evidence suggests pivotal role of IL-22 in instigation of various cancers due to its pro-inflammatory and tissue repairing activity. In this review, we summarize how healing effects of IL-22, when executed in an uncontrollable fashion can lead to carcinogenesis.

Comparative analysis of Bacterial Vaginosis microbiota among pregnant and non-pregnant females and isolation of phages against Enterococcus faecalis, Enterococcus faecium, and Shigella flexneri strains.

BACKGROUND AND OBJECTIVES: Bacterial vaginosis (BV) is the most common vaginal infection in women of reproductive age. It shifts the paradigms of the vagina from healthy, beneficial microbiota to facultative and strict anaerobes. BV remains one of the most arduous and controversial challenges in modern-day clinical microbiology because of its high prevalence and relapse rates. A lot of research has been carried out on it. Still, its etiology is unknown, which gave this infection global importance. The current study was designed to investigate and compare the microbiota of pregnant and non-pregnant females suffering from BV, and phages were isolated against BV microbiota. MATERIAL AND METHODS: The samples were collected from the vagina by using a speculum, and swabs were streaked on different media to isolate bacteria. The microbiological analysis was performed by microscopy, biochemical testing, and antibiotic susceptibility was determined by using Metronidazole and Clindamycin. Furthermore, the phages were isolated and characterized against BV strains. RESULTS AND CONCLUSION: The Gram staining showed high prevalence of Staphylococcus (36% vs. 33%), followed by Streptococcus (31% vs. 14%) and Enterococcus (7% vs. 14%) in non-pregnant and pregnant females' respectively. However, the exception was observed in non-pregnant BV positive females, who had Shigella flexneri in their samples. The antibiotic sensitivity showed Metronidazole was resistant against all BV microbiota, and Clindamycin showed susceptibility against 3 strains. Phages were isolated against three bacterial strains, i.e. E. faecalis, E. faecium, and S. flexneri. Bacterial reduction assay showed bacterial growth decreases in the presence of phage suspension, pH stability showed phages' maximum lytic activity at pH 7 for E. faecalis and E. faecium and pH 9 for S. flexneri. However, the thermal stability showed phages' highest lytic activity at 55 °C for E. faecalis, 70 °C for E. faecium, and 40 °C for S. flexneri. Phage genome isolation showed that all phages nucleic acid was DNA in nature and between 15 and 20kbp. SEM analysis showed they were circular in shape and might belong to the Podoviridae family. This study provides an understanding of pathogens involved in BV and helps the doctors to treat the patients accordingly. Furthermore, this study showed that Bacterial Vaginosis and BV secondary bacteria have associations. BV secondary microbiota is also involved in the pathogenesis of this infection, whereas bacteriophage therapy has the potential to be used as an alternative treatment to antibiotics.

Efficacy and Safety of Generic Sofosbuvir Plus Daclatasvir and Sofosbuvir/Velpatasvir in HCV Genotype 3-Infected Patients: Real-World Outcomes From Pakistan.

BACKGROUND: Direct-acting antivirals (DAAs) therapeutic regimens are highly effective against chronic hepatitis C virus (HCV) infection. However, HCV patients with genotype 3 (GT3) respond in a suboptimal way. This study aims to identify which of the DAAs-based therapeutic regimens are the best option for GT3. METHODS: Multiple governments and private tertiary care hospitals were involved in this real-life study of HCV-GT3 patients treated with DAAs. The efficacy and safety of generic sofosbuvir+daclatasvir±ribavirin (SOF+DCV±RBV) and sofosbuvir/velpatasvir±ribavirin (SOF/VEL±RBV) were assessed under the National Hepatitis C Program of Pakistan. RESULTS: Out of 1,388 participants, 70% of patients received SOF+DCV in government tertiary care hospitals and 30% received SOF/VEL in private tertiary care hospitals. The overall sustained virological responses (SVR) was 95.5%. The SVR rates at 12 weeks were comparable between SOF+DCV (94.4%) and SOF/VEL (94.7%) in chronic HCV patients. However, The SVR rates at 24 weeks were high in cirrhotic patients treated with SOF/VEL+RBV (88%) then SOF+DCV+RBV (83%). Non-responders were high in SOF-DCV than SOF-VEL (4.1 vs 3.8%, P = 0.05) regimen. In multivariate models, the significant predictors of non-SVR were age >60 years (odds ratio [OR] 4.46; 95% CI, 2.35-8.46, P = <0.001) and cirrhosis (OR 53.91; 95% CI, 26.49-109.6, P = <0.001). Skin rash (51 vs 44%) and oral ulcers (45 vs 40%) were high in patients receiving SOF-DCV then SOF-VEL. CONCLUSIONS: Overall, the generic SOF+DCV ±RBV and SOF/VEL ± RBV achieved equally high SVR12 rates. However, SOF/VEL+RBV achieved a high SVR rate in cirrhotic patients then SOF+DCV+RBV. Old age and cirrhosis were significant predictors of reduced odds of SVR regardless of the regimen. Furthermore, the regimens were well tolerated in chronic HCV patients.

Induction of Profibrotic Microenvironment via TLR4 MyD88-Dependent and -Independent Inflammatory Signaling in Chronic Hepatitis C Virus Infection.

Hepatitis C virus (HCV) is a well-known pathogen to establish chronic infection leading to end-stage liver disease. The destruction of liver tissues takes its roots under chronic inflammation and proinflammatory signaling in liver microenvironment. The viral proteins interact with certain pattern recognition receptors, including Toll-like receptors, activating the innate immune system to clear the virus. HCV achieves immune evasion through other mechanisms and induce a continuous inflammatory microenvironment via Kupffer cells and Hepatic Stellate cells. This promotes disease progression. The current study aims to elucidate that the role of Toll-like receptor 4 (TLR4) induced innate immune response in chronic inflammation in patients chronically infected with HCV. For this purpose, changes in downstream signaling cascade of TLR4 during chronic HCV infection using peripheral blood mononuclear cells of chronic HCV patients were studied. We found significant increase in expression levels of proinflammatory and profibrotic genes induced by TLR4 Myeloid differentiation factor 88 (MyD88)-dependent pathway between treatment naive and healthy controls, while no significant difference between the expressions of genes involved in TLR4 signaling was found between treatment responders and healthy controls. Interestingly, both TLR4 MyD88-dependent and -independent pathways were found to be operational in nonresponders to interferon treatment. This further strengthens the involvement of innate immune signaling as a leading factor in HCV-mediated liver disease progression and the role of TLR4 MyD88-dependent and -independent pathway in ensuring the conditions for chronic inflammation.

Tumor educated platelets, a promising source for early detection of hepatocellular carcinoma: Liquid biopsy an alternative approach to tissue biopsy.

PURPOSE: Liver cancer is considered to be the sixth deadliest cancer worldwide. Hepatocellular carcinoma (HCC) is known to be the most prevalent type of liver cancer. The number of deaths due to HCC reported per year is on a constant rise especially in lesser developed countries. There are several contributing factors to this rise in number. Among other contributing factors is the late diagnosis of HCC. Patients are usually diagnosed when the disease reaches its advance stage. The present study was conducted with total 30 samples. It was designed for investigating the potential of TGF-ß, NF-κß, VEGF, AKT and PI3K as RNA based biomarkers in tumor educated platelets for early detection of HCC. RESULTS: The results obtained from the transcriptional analysis revealed a significant high expression of TGF-ß, NF-κß, VEGF by 2.48, 2.35 and 2.78 folds respectively in comparison to the control. On the other hand, a decrease in expression by 0.6 and 0.65 folds was observed in AKT and PI3K respectively in comparison to controls. Although all selected RNA biomarkers showed promising potential to detect HCC however, AKT and PI3K were better able to detect early stage HCC. CONCLUSIONS: The results obtained clearly indicate the increased expression of TGF-ß, NF-κß, VEGF in HCC patients. All these biomarkers are previously known for cancer initiation, progression and metastasis. The significant decrease in expression of AKT and PI3K in HCC patients needs further investigation. All the selected RNA biomarkers can be used for detection of HCC as they were able to distinguish HCC patients from controls successfully with AKT and PI3K showing better potential to detect early stage HCC. However, translational analysis for all these RNA biomarkers should be performed to gain further evidence for the ability of these biomarkers to be used for early HCC detection.

Direct-acting antiviral agents in the treatment of chronic hepatitis C-Real-life experience from clinical practices in Pakistan.

This study aims to evaluate the clinical effectiveness in terms of sustained virological response (SVR), predictors of SVR and safety of available second-generation generic direct-acting antivirals in Pakistani chronic hepatitis C patients. This is a retrospective study conducted in multiple centers of Pakistan from January 2015 to January 2019. The samples include patients infected with chronic hepatitis C virus, regardless of virus genotype, cirrhosis, or prior treatment. A total of 993 patients were included in the present study, with the majority receiving sofosbuvir with daclatasvir (95%), sofosbuvir with daclatasvir and ribavirin (4%), and sofosbuvir with ribavirin (1%). There were 96% cases of chronic hepatitis, 3% cases compensated cirrhosis, and 1% cases of decompensated cirrhosis. Genotype 3 (99.6%) was the most common genotype. Overall SVR after 12 weeks was 98% for all treatment regimens. High SVR12 was observed with sofosbuvir in combination with daclatasvir (98.5%), then sofosbuvir in combination with daclatasvir and ribavirin (90.2%) and sofosbuvir in combination with ribavirin (75%). SVR rates were high in chronic hepatitis C patients (98.2%) as compared with cirrhotic patients (92.1%) and it was high in treatment-naive (98.8%) then interferon experienced patients (90.1%). In multivariate binary logistic regression analysis, patients' education status, treatment strategy, viral load, and alanine aminotransferase had a statistically significant association with SVR at 12 weeks. No major adverse events occurred which required treatment discontinuation. Generic oral direct acting antiviralss (sofosbuvir with daclatasvir) achieved higher SVR12 rates and were well tolerated in this large real-world cohort of genotype 3 infected patients.