Prevalence of Hepatitis C Viral Infection among Diabetes Mellitus Patients in Qassim Region, Saudi Arabia

The worldwide prevalence of Diabetes Mellitus (DM) associated with Hepatitis C Virus (HCV) infection are reported with higher rates of morbidity and mortality. The frequency of HCV is approximately 3-4 million cases each year and in parallel the incidence of DM is increasing alarmingly. World Health Organization (WHO) has specified that DM will be the 7th leading cause of mortality by 2030. The increasing association between HCV and DM has been indicated by some significant reports recently. HCV infection leads to hepatic steatosis and rapid insulin resistance, which in turn upsurges the risk factors for hepatic fibrosis and hepatocellular carcinoma. This study is designed to examine the association between HCV and DM, and different risk factors associated with HCV infection in Qassim region, Kingdom of Saudi Arabia (KSA). A total of 634 blood samples were obtained from diabetic and non-diabetic patients. These blood samples were first screened for HCV infection by enzyme-linked immunosorbent assay (ELISA) and positive samples were again confirmed by TaqMan HCV quantitative test and the viral load in different samples was estimated. The HCV prevalence was identified as 2.5% in diabetic patients with a positive association between HCV and DM (RR= 1.24, OR= 1.77) which is not significant statistically. However, the HCV prevalence among diabetic females was significantly different from males (p<0.05). The behavioural factors had no significant impact to acquire HCV infection. This study indicated a positive association between HCV and DM. Gender was an association factor in the HCV and DM status. Further studies with larger sample size is significant to properly assess the temporal relationship between HCV and DM

Blue biotechnology: Computational screening of sarcophyton cembranoid diterpenes for SARS-CoV-2 main protease inhibition

The coronavirus pandemic has affected more than 150 million people, while over 3.25 million people have died from the coronavirus disease 2019 (COVID-19). As there are no established therapies for COVID-19 treatment, drugs that inhibit viral replication are a promising target; specifically, the main protease (Mpro) that process CoV-encoded polyproteins serves as an Achilles heel for assembly of replication-transcription machinery as well as down-stream viral replication. In the search for potential antiviral drugs that target Mpro, a series of cembranoid diterpenes from the biologically active soft-coral genus Sarcophyton have been examined as SARS-CoV-2 Mpro inhibitors. Over 360 metabolites from the genus were screened using molecular docking calculations. Promising diterpenes were further characterized by molecular dynamics (MD) simulations based on molecular mechanics-generalized Born surface area (MM-GBSA) binding energy calculations. According to in silico calculations, five cembranoid diterpenes manifested adequate binding affinities as Mpro inhibitors with ΔGbinding \u3c -33.0 kcal/mol. Binding energy and structural analyses of the most potent Sarcophyton inhibitor, bislatumlide A (340), was compared to darunavir, an HIV protease inhibitor that has been recently subjected to clinical-trial as an anti-COVID-19 drug. In silico analysis indicates that 340 has a higher binding affinity against Mpro than darunavir with ΔGbinding values of -43.8 and -34.8 kcal/mol, respectively throughout 100 ns MD simulations. Drug-likeness calculations revealed robust bioavailability and protein-protein interactions were identified for 340; biochemical signaling genes included ACE, MAPK14 and ESR1 as identified based on a STRING database. Pathway enrichment analysis combined with reactome mining revealed that 340 has the capability to re-modulate the p38 MAPK pathway hijacked by SARS-CoV-2 and antagonize injurious effects. These findings justify further in vivo and in vitro testing of 340 as an antiviral agent against SARS-CoV-2

A Comprehensive Computational Screening of Phytochemicals Derived from Saudi Medicinal Plants against Human CC Chemokine Receptor 7 to Identify Potential Anti-Cancer Therapeutics

Homeostatic trafficking of immune cells by CC chemokine receptor 7 (CCR7) keeps immune responses and tolerance in a balance. The involvement of this protein in lymph node metastasis in cancer marks CCR7 as a penitential drug target. Using the crystal structure of CCR7, herein, a comprehensive virtual screening study is presented to filter novel strong CCR7 binding phytochemicals from Saudi medicinal plants that have a higher binding affinity for the intracellular allosteric binding pocket. By doing so, three small natural molecules named as Hit-1 (1,8,10-trihydroxy-3-methoxy-6-methylanthracen-9(4H)-one), Hit-2 (4-(3,4-dimethoxybenzyl)-3-(4-hydroxy-3-methoxybenzyl)dihydrofuran-2(3H)-one), and Hit-3 (10-methyl-12,13-dihydro-[1,2]dioxolo[3,4,5-de]furo[3,2-g]isochromeno[4,3-b]chromen-8-ol) are predicted showing strong binding potential for the CC chemokine receptor 7 allosteric pocket. During molecular dynamics simulations, the compounds were observed in the formation of several chemical bonding of short bond distances. Additionally, the molecules remained in strong contact with the active pocket residues and experienced small conformation changes that seemed to be mediated by the CCR7 loops to properly engage the ligands. Two types of binding energy methods (MM/GBPBSA and WaterSwap) were additionally applied to further validate docking and simulation findings. Both analyses complement the good affinity of compounds for CCR7, the electrostatic and van der Waals energies being the most dominant in intermolecular interactions. The active pocket residue’s role in compounds binding was further evaluated via alanine scanning, which highlighted their importance in natural compounds binding. Additionally, the compounds fulfilled all drug-like rules: Lipinski, Ghose, Veber, Egan, and Muegge passed many safety parameters, making them excellent anti-cancer candidates for experimental testing

The Effect of Liposomal Diallyl Disulfide and Oxaliplatin on Proliferation of Colorectal Cancer Cells: In Vitro and In Silico Analysis

Diallyl disulfide (DADS) is one of the main bioactive organosulfur compounds of garlic, and its potential against various cancer models has been demonstrated. The poor solubility of DADS in aqueous solutions limits its uses in clinical application. The present study aimed to develop a novel formulation of DADS to increase its bioavailability and therapeutic potential and evaluate its role in combination with oxaliplatin (OXA) in the colorectal cancer system. We prepared and characterized PEGylated, DADS (DCPDD), and OXA (DCPDO) liposomes. The anticancer potential of these formulations was then evaluated in HCT116 and RKO colon cancer cells by different cellular assays. Further, a molecular docking-based computational analysis was conducted to determine the probable binding interactions of DADS and OXA. The results revealed the size of the DCPDD and DCPDO to be 114.46 nm (95% EE) and 149.45 nm (54% EE), respectively. They increased the sensitivity of the cells and reduced the IC50 several folds, while the combinations of them showed a synergistic effect and induced apoptosis by 55% in the cells. The molecular docking data projected several possible targets of DADS and OXA that could be evaluated more precisely by these novel formulations in detail. This study will direct the usage of DCPDD to augment the therapeutic potential of DCPDO against colon cancer in clinical settings

Assessment of UVR-Induced DNA Damage and Repair in Nuclear Genome versus Mitochondrial Genome

DNA is a key molecular-target for the deleterious effects of ultraviolet radiation (UVR). Cells contain two types of DNA: nuclear DNA (nDNA) and mitochondrial DNA (mtDNA) and UVR induces various types of damage in the both DNAs, notably CPDs and 8-oxodG. The aim of this thesis is to examine UVR induced DNA damage formation and repair in nDNA and mtDNA and to determine which is the most important genomic target with respect to cell killing in vitro using HaCaT calls as models of human skin. The cell viability data showed that UVB induces significant cell death, which increased over 48 h. SSR-exposure also showed significant levels of cell death after 24 h but with evidence of significant survival after 48 h. Alkaline modified comet assay data showed that CPDs and 8-oxodG were significantly produced in HaCaT cells exposed to UVB and SSR, with CPDs being formed in a greater yields and there being no significant repair of CPDs over 48 h post-exposure to UVB. However, HaCaT cells irradiated with SSR showed significant repair of both CPD and 8-oxodG over 48 h. QPCR data showed that UVB and SSR induced similar profiles of damage in both nDNA and mtDNA; despite the induced damage levels being higher with UVB. The data also showed that nDNA is the main target for UVR in HaCaT cells exposed to UVB and SSR. The UVB-induced QPCR-detectable DNA damage in nDNA and mtDNA was not fully repaired, with a significant level of DNA damage remaining at 48 h, however, there was significant repair of the induced-damage in nDNA post-exposure to SSR (correlating with survival/re-growth), whereas the damage to mtDNA was not fully repaired. The greater lethality of UVB is probably due to more the damage induced and poorer repair (notably of CPD) in nuclear DNA following UVB exposure. Whereas the proficient repair of SSR-induced CPD in nDNA probably dictates survival following SSR exposure – as there was still a notable level of residual damage in mtDNA post-SSR exposure. However, nDNA is the main target for UVR causing DNA damage and may lead to mutations, which increase the risk of skin cancer development

Therapeutic Effect of Bilsaan, Sambucus nigra Stem Exudate, on the OVA-Induced Allergic Asthma in Mice

Asthma is characterized by the elevated level of Th2 immune responses, oxidative stress, and airway inflammation. Bilsaan, an exudate from the stem of Sambucus nigra, has been traditionally used in the treatment of various ailments in Saudi Arabia. Here, we investigated the therapeutic potential of Bilsaan against ovalbumin- (OVA-) induced allergic asthma in a mouse model. In order to induce allergic asthma, mice were intraperitoneally injected with alum-emulsified-OVA (20 μg/mouse) on days 0, 14, and 21 that is followed by an intranasal OVA exposure from days 22 to 30. During this time, mice were orally administered with Bilsaan at the doses of 5, 10, and 25 mg/kg. The numbers of total and differential inflammatory cells and the levels of Th2 cytokines (IL-4, IL-5, and IL-13) and IgE were determined in bronchoalveolar lavage fluid (BALF). Moreover, the therapeutic effect of Bilsaan was also assessed to analyze the oxidative stress and inflammatory changes in the lung tissues. The results demonstrated that Bilsaan treatment significantly reduced the total and differential inflammatory cell count in the BALF. The BALF from the mice treated with Bilsaan showed significantly lower levels of IL-4, IL-5, IL-13, and IgE. Interestingly, a similar pattern was observed in IL-4, IL-5, and IL-13 secreted by OVA-sensitized splenocytes from the mice of various groups. Bilsaan treatment alleviated the status of oxidative stress by modulating malondialdehyde (MDA), superoxide dismutase (SOD), and catalase levels in the lung. Moreover, Bilsaan treatment reduced the infiltration of inflammatory cells, thickening of alveolar wall, and congestion in the lung tissues. The findings of the present study demonstrated an antiasthmatic effect of Bilsaan through the modulation of Th2 immune responses, inflammation, and the oxidative stress

Design of a Multi-Epitopes Based Chimeric Vaccine against Enterobacter cloacae Using Pan-Genome and Reverse Vaccinology Approaches

Enterobacter cloacae (EC) is a significant emerging pathogen that is occasionally associated with lung infection, surgical site infection, urinary infection, sepsis, and outbreaks in neonatal intensive care units. In light of the fact that there is currently no approved vaccine or therapeutic option for the treatment of EC, the current study was developed to concentrate on applications based on modern computational approaches to design a multi-epitope-based E. cloacae peptide vaccine (MEBEPV) expressing the antigenic determinants prioritized from the EC genome. Integrated computational analyses identified two potential protein targets (phosphoporin protein-PhoE and putative outer-membrane porin protein) for further exploration on the basis of pangenome subtractive proteomics and immunoinformatic in-depth examination of the core proteomes. Then, a multi-epitope peptide vaccine was designed, which comprised shortlisted epitopes that were capable of eliciting both innate and adaptive immunity, as well as the cholera toxin&rsquo;s B-subunit, which was used as an adjuvant in the vaccine formulation. To ensure maximum expression, the vaccine&rsquo;s 3D structure was developed and the loop was refined, improving the stability by disulfide engineering, and the physicochemical characteristics of the recombinant vaccine sequence were found to be ideal for both in vitro and in vivo experimentation. Blind docking was then used for the prediction of the MEBEPV predominant blinding mode with MHCI, MHCII, and TLR3 innate immune receptors, with lowest global energy of &minus;18.64 kJ/mol, &minus;48.25 kJ/mol, and &minus;5.20 kJ/mol for MHC-I, MHC-II, and TLR-4, respectively, with docked complexes considered for simulation. In MD and MMGBSA investigations, the docked models of MEBEPV-TLR3, MEBEPV-MHCI, and MEBEPV-MHCII were found to be stable during the course of the simulation. MM-GBSA analysis calculated &minus;122.17 total net binding free energies for the TLR3-vaccine complex, &minus;125.4 for the MHC I-vaccine complex, and &minus;187.94 for the MHC II-vaccine complex. Next, MM-PBSA analysis calculated &minus;115.63 binding free energy for the TLR3-vaccine complex, &minus;118.19 for the MHC I-vaccine complex, and &minus;184.61 for the MHC II-vaccine complex. When the vaccine was tested in silico, researchers discovered that it was capable of inducing both types of immune responses (cell mediated and humoral) at the same time. Even though the suggested MEBEPV has the potential to be a powerful contender against E. cloacae-associated illnesses, further testing in the laboratory will be required before it can be declared safe and immunogenic

Hepatoprotective Effects of Garlic Extract against Carbon Tetrachloride (CCl4)-Induced Liver Injury via Modulation of Antioxidant, Anti-Inflammatory Activities and Hepatocyte Architecture

The current study aims to explore the hepatoprotective mechanisms of garlic extract through in vivo and in vitro assays. The in vitro investigation of antioxidant and anti-inflammatory potential showed maximum 67.5% of free radical scavenging and 71.36% albumin denaturation inhibition by 600 &mu;g/mL garlic extract. To explore the hepatoprotective activity by in vivo experiments, the animals were orally intoxicated with 150 &mu;L of CCl4 (1:1 v/v in olive oil) and treated with garlic extract (75 mg/kg b.w.) 3 times/week, for eight successive weeks. The administration of garlic extract significantly ameliorated CCl4 induced increment in amounts of serum Alanine aminotransferase (ALT), Alkaline phosphatase (ALP) and Aspartate transaminaseas (106.7, 116.3, 136.4 U/L) as compared to disease control which showed increased level (140.5, 156.2, 187.6 U/L). Besides, significant reduction of Superoxide dismutase (SOD), Glutathione peroxidases (GPx), and Glutathione (GSH) (29.3, 48.4, and 25.9 U/mg protein) was noticed in CCl4 induced animals, respectively. Likewise, garlic extract treatment facilitated a significant increment in all tested antioxidant enzymes levels (41.6, 63.3, and 32.5 U/mg protein), respectively. Additionally, Tumor necrosis factor⍺ (TNF-⍺), C-reactive protein (CRP), Interleukin-1&beta; (IL-1&beta;), Interleukin 6 (IL-6) and ICAM-1 (Intercellular Adhesion Molecule 1) level (63.79, 580.2, 18.3, 63.74 and 148.4 pg/mL) were increased significantly in CCl4-induced group, while garlic extract treatment decreased these pro inflammatory marker levels (40.24, 460.4, 15.4, 45.14, and 125.3 pg/mL). The animals exposed to CCl4 showed various types of alterations like lymphocytes infiltration, edema and congestion, while the animals treated with garlic extract plus CCl4 showed amelioration of the hepatocytes architectures. Thus, our finding advocates that the consumption of garlic can be a potential therapeutic remedy in the inhibition of liver ailments

6-Gingerol, a Major Ingredient of Ginger Attenuates Diethylnitrosamine-Induced Liver Injury in Rats through the Modulation of Oxidative Stress and Anti-Inflammatory Activity

Diethylnitrosamine (DEN) is a well-known hepatocarcinogen, and its oral administration causes severe liver damage including cancer. DEN induces the pathogenesis of the liver through reactive oxygen species mediated inflammation and modulation of various biological activities. 6-Gingerol, a major component of ginger, is reported to prevent liver diseases by reducing the oxidative stress and proinflammatory mediators. The present study investigated the hepatoprotective effects of 6-gingerol through the measurement of oxidative stress, anti-inflammatory markers, liver function enzyme parameter, and histopathological analysis. The rats were randomly divided into four groups as the control, DEN treated (50 mg/kg b.w.), DEN+6-gingerol (each 50 mg/kg b.w.), and 6-gingerol only. To evaluate the hepatoprotective effects, liver function enzymes (ALT, AST, and ALP), oxidative stress markers (SOD, GSH, GST, and TAC), lipid peroxidation, inflammatory markers (CRP, TNF-α, IL-6, and ICAM1), haematoxylin and eosin staining, Sirius red staining, immunohistochemistry, and electron microscopy were performed. The results showed a significant increase in liver function enzymes, oxidative stress, and inflammatory markers in the DEN-treated group as compared to the control group. Besides this, altered architecture of hepatocytes (infiltration of inflammatory cells, congestion, blood vessel dilation, and edema), abundant collagen fiber and organelle structures like distorted shaped and swollen mitochondria, and broken endoplasmic reticulum were noticed. The administration of 6-gingerol significantly ameliorated the biochemical and histopathological changes. The increased expression of TNF-α protein was noticed in the DEN-treated group whereas the administration of 6-gingerol significantly decreased the expression of this protein. Based on these findings, it can be suggested that 6-gingerol may be an alternative therapy for the prevention and treatment of liver diseases

Comprehensive Integrative Analysis Reveals the Association of KLF4 with Macrophage Infiltration and Polarization in Lung Cancer Microenvironment

Macrophage polarization and infiltration to the tumor microenvironment (TME) is a critical determining factor for tumor progression. Macrophages are polarized into two states—M1 (pro-inflammatory, anti-tumorigenic and stimulated by LPS or IFN-γ) and M2 (anti-inflammatory pro-tumorigenic and stimulated by IL-4) phenotypes. Specifically, M2 macrophages enhance tumor cell growth and survival. Recent evidences suggest the pivotal role of microRNAs in macrophage polarization during the development of Non-small cell lung cancer (NSCLC), thus proposing a new therapeutic option to target lung cancer. In silico analysis determined cogent upregulation of KLF4, downregulation of IL-1β and miR-34a-5p in NSCLC tissues, consequently worsening the overall survival of NSCLC patients. We observed a significant association of KLF4 with macrophage infiltration and polarization in NSCLC. We found that KLF4 is critically implicated in M2 polarization of macrophages, which, in turn, promotes tumorigenesis. KLF4 expression correlated with miR-34a-5p and IL-1β in a feed-forward loop (FFL), both of which are implicated in immune regulation. Mechanistic overexpression of miR-34a-5p in macrophages (IL-4 stimulated) inhibits KLF4, along with downregulation of ARG1, REL-1MB (M2 macrophage specific markers), and upregulation of IL-1β, IL-6, (M1 macrophage specific markers), demonstrating macrophage polarization switch from M2 to M1 phenotype. Moreover, co-culture of these macrophages with NSCLC cells reduces their proliferation, wound healing, clonogenic capacity and enhanced NO-mediated apoptosis. Further, transfection of miR-34a-5p in NSCLC cells, also degrades KLF4, but enhances the expression of KLF4 regulated genes—IL-1β, IL-6 (pro-inflammatory mediators), which is further enhanced upon co-culture with IL-4 stimulated macrophages. Additionally, we observed a significant increase in i-NOS/NO content upon co-culture, suggesting polarization reversion of macrophages from M2 to M1, and eventually leading to anti-tumor effects. Our findings thus show a significant role of KLF4 in tumorigenesis and TAM polarization of NSCLC. However, miR-34a-5p mediated targeting of these molecular networks will provide a better therapeutic intervention for NSCLC