Probiotic-derived silver nanoparticles target mTOR/MMP-9/BCL-2/dependent AMPK activation for hepatic cancer treatment.

Recent advances in nanotechnology have offered novel ways to combat cancer. By utilizing the reducing capabilities of Lactobacillus acidophilus, silver nanoparticles (AgNPs) are synthesized. The anti-cancer properties of AgNPs have been demonstrated in previous studies against several cancer cell lines; it has been hypothesized that these compounds might inhibit AMPK/mTOR signalling and BCL-2 expression. Consequently, the current research used both in vitro and in silico approaches to study whether Lactobacillus acidophilus AgNPs could inhibit cell proliferation autophagy and promote apoptosis in HepG2 cells. The isolated strain was identified as Lactobacillus acidophilus strain RBIM based on 16 s rRNA gene analysis. Based on our research findings, it has been observed that this particular strain can generate increased quantities of AgNPs when subjected to optimal growing conditions. The presence of silanols, carboxylates, phosphonates, and siloxanes on the surface of AgNPs was confirmed using FTIR analysis. AgNPs were configured using UV-visible spectroscopy at 425 nm. In contrast, it was observed that apoptotic cells exhibited orange-coloured bodies due to cellular shrinkage and blebbing initiated by AgNP treatment, compared to non-apoptotic cells. It is worth mentioning that AgNPs exhibited remarkable selectivity in inducing cell death, specifically in HepG2 cells, unlike normal WI-38 cells. The half-maximum inhibitory concentration (IC50) values for HepG2 and WI-38 cells were 4.217 µg/ml and 154.1 µg/ml, respectively. AgNPs induce an upregulation in the synthesis of inflammation-associated cytokines, including (TNF-α and IL-33), within HepG2 cells. AgNPs co-treatment led to higher glutathione levels and activating pro-autophagic genes such as AMPK.Additionally, it resulted in the suppression of mTOR, MMP-9, BCL-2, and α-SMA gene expression. The docking experiments suggest that the binding of AgNPs to the active site of the AMPK enzyme leads to inhibiting its activity. The inhibition of AMPK ultimately results in the suppression of the mechanistic mTOR and triggers apoptosis in HepG2 cells. In conclusion, the results of our study indicate that the utilization of AgNPs may represent a viable strategy for the eradication of liver cancerous cells through the activation of apoptosis and the enhancement of immune system reactions.

Anti-Tumor Potential of Gymnema sylvestre Saponin Rich Fraction on In Vitro Breast Cancer Cell Lines and In Vivo Tumor-Bearing Mouse Models.

Gymnema sylvestre (GS) is a perennial woody vine native to tropical Asia, China, the Arabian Peninsula, Africa and Australia. GS has been used as a medicinal plant with potential anti-microbial, anti-inflammatory and anti-oxidant properties. This study was conceptualized to evaluate the cytotoxicity potential of Gymnema sylvestre saponin rich fraction (GSSRF) on breast cancer cell lines (MCF-7 and MDA-MB-468) by SRB assay. The anti-tumor activity of GSSRF was assessed in tumor-bearing Elrich ascites carcinoma (EAC) and Dalton's lymphoma ascites (DLA) mouse models. The anti-oxidant potential of GSSRF was assessed by DPPH radical scavenging assay. The acute toxicity of GSSRF was carried out according to OECD guideline 425. The yield of GSSRF was around 1.4% and the presence of saponin content in GSSRF was confirmed by qualitative and Fourier transform infrared spectroscopic (FTIR) analysis. The in vitro cytotoxic effects of GSSRF on breast cancer cell lines were promising and found to be dose-dependent. An acute toxicity study of GSSRF was found to be safe at 2000 mg/kg body weight. GSSRF treatment has shown a significant increase in the body weight and the life span of EAC-bearing mice in a dose-dependent manner when compared with the control group. In the solid tumor model, the doses of 100 and 200 mg/kg body weight per day have shown about 46.70% and 60.80% reduction in tumor weight and controlled the tumor weight until the 30th day when compared with the control group. The activity of GSSRF in both models was similar to the cisplatin, a standard anticancer agent used in the study. Together, these results open the door for detailed investigations of anti-tumor potentials of GSSRF in specific tumor models, mechanistic studies and clinical trials leading to promising novel therapeutics for cancer therapy.

Identification of compelling inhibitors of human norovirus 3CL protease to combat gastroenteritis: A structure-based virtual screening and molecular dynamics study.

Human noroviruses (NV) are the most prevalent cause of sporadic and pandemic acute gastroenteritis. NV infections cause substantial morbidity and death globally, especially amongst the aged, immunocompromised individuals, and children. There are presently no authorized NV vaccines, small-molecule therapies, or prophylactics for humans. NV 3 C L protease (3CLP) has been identified as a promising therapeutic target for anti-NV drug development. Herein, we employed a structure-based virtual screening method to screen a library of 700 antiviral compounds against the active site residues of 3CLP. We report three compounds, Sorafenib, YM201636, and LDC4297, that were revealed to have a higher binding energy (BE) value with 3CLP than the control (Dipeptidyl inhibitor 7) following a sequential screening, in-depth molecular docking and visualization, physicochemical and pharmacological property analysis, and molecular dynamics (MD) study. Sorafenib, YM201636, and LDC4297 had BEs of -11.67, -10.34, and -9.78 kcal/mol with 3CLP, respectively, while control had a BE of -6.38 kcal/mol. Furthermore, MD simulations of the two best compounds and control were used to further optimize the interactions, and a 100 ns MD simulation revealed that they form stable complexes with 3CLP. The estimated physicochemical, drug-like, and ADMET properties of these hits suggest that they might be employed as 3CLP inhibitors in the management of gastroenteritis. However, wet lab tests are a prerequisite to optimize them as NV 3CLP inhibitors.

Case Report: Biallelic Variant in the tRNA Methyltransferase Domain of the AlkB Homolog 8 Causes Syndromic Intellectual Disability.

Intellectual disability (ID) has become very common and is an extremely heterogeneous disorder, where the patients face many challenges with deficits in intellectual functioning and adaptive behaviors. A single affected family revealed severe disease phenotypes such as ID, developmental delay, dysmorphic facial features, postaxial polydactyly type B, and speech impairment. DNA of a single affected individual was directly subjected to whole exome sequencing (WES), followed by Sanger sequencing. Data analysis revealed a novel biallelic missense variant (c.1511G>C; p.(Trp504Ser)) in the ALKBH8 gene, which plays a significant role in tRNA modifications. Our finding adds another variant to the growing list of ALKBH8-associated tRNA modifications causing ID and additional phenotypic manifestations. The present study depicts the key role of the genes associated with tRNA modifications, such as ALKBH8, in the development and pathophysiology of the human brain.

Targeting Cytotoxin-Associated Antigen A, a Virulent Factor of Helicobacter pylori-Associated Gastric Cancer: Structure-Based In Silico Screening of Natural Compounds.

Gastric cancer is the fifth most frequent cancer and the third major cause of mortality worldwide. Helicobacter pylori, a bacterial infection linked with GC, injects the cytotoxin-associated antigen A (CagA; an oncoprotein) into host cells. When the phosphorylated CagA protein enters the cell, it attaches to other cellular components, interfering with normal cellular signaling pathways. CagA plays an important role in the progression of GC by interacting with phosphatidylserine of the host cell membrane. Therefore, disrupting the CagA-phosphatidylserine connection using small molecules appears to be a promising therapeutic approach. In this report, we screened the natural compounds from ZINC database against the CagA protein using the bioinformatics tools. Hits were initially chosen based on their physicochemical, absorption, distribution, metabolism, excretion, and toxicity (ADMET) characteristics, as well as other drug-like characteristics. To locate safe and effective hits, the PAINS filter, binding affinities estimation, and interaction analysis were used. Three compounds with high binding affinity and specificity for the CagA binding pocket were discovered. The final hits, ZINC153731, ZINC69482055, and ZINC164387, were found to bind strongly with CagA protein, with binding energies of -11.53, -10.67, and -9.21 kcal/mol, respectively, which were higher than that of the control compound (-7.25 kcal/mol). Further, based on binding affinity and interaction pattern, two leads (ZINC153731, ZINC69482055) were chosen for molecular dynamics (MD) simulation analysis. MD results showed that they displayed stability in their vicinity at 100 ns. This study suggested that these compounds could be used as possible inhibitors of CagA protein in the fight against GC. However, additional benchwork tests are required to validate them as CagA protein inhibitors.

In vitro and in silico characterization of alkaline serine protease from Bacillus subtilis D9 recovered from Saudi Arabia.

In this study, we have isolated and characterized proteolytic soil bacteria and their alkaline protease. Based on 16S rRNA sequence analysis, 12 isolates with the highest protease activity were classified as B. subtilis and B. cereus groups. B. subtilis D9 isolate showing the highest protease activity was selected for in vitro and in silico analysis for its ِِAKD9 protease. The enzyme has a molecular mass of 48 kDa, exhibiting optimal activity at 50 °C pH 9.5, and showed high stability till 65 °C and pH 8-11 for 1 h. Fe3+ stimulated, but Zn2+ and Hg2+ strongly inhibited the protease activity. Also, the maximum inhibition with PMSF indicated serine protease-type of AKD9 protease. AkD9 alkaline serine protease gene showed high sequence similarity and close phylogenetic relationship with AprX serine protease of B. subtilis isolates. Functional prediction of AKD9 resulted in the detection of subtilase domain, peptidase_S8 family, and subtilase active sites. Moreover, prediction of physicochemical properties indicated that AKD9 serine protease is hydrophilic, thermostable, and alkali-halo stable. Secondary structure prediction revealed the dominance of the coils enhances AKD9 activity and stability under saline and alkaline conditions. Based on molecular docking, AKD9 showed very promising binding affinities towards casein substrate with expected intrinsic proteolytic activities matching our obtained in vitro results. In conclusion, AKD9 alkaline serine protease seems to be a significant candidate for industrial applications because of its stability, hydrophilicity, enhanced thermostability, and alkali-halo stability.

Histological Studies on a Newly Isolated Bacillus subtilis D10 Protease in the Debridement of Burn Wound Eschars Using Mouse Model.

BACKGROUND: Proteases are among the most important industrial enzymes, playing a critical role in the physiological, biochemical, and regulatory processes of all living organisms. This study evaluated the histological effects of a Bacillus subtilis D10 protease in combination with the antibacterial ointment silver sulfadiazine (SSD) on the burned skin of mice. MATERIALS AND METHODS: The bacterial proteolytic enzyme was produced and purified through DEAE-Sepharose CL-6B and Sephadex G-100 FF. The in vitro protease specificity was then determined. The dorsal skin of albino mice was burned with 80% HCl solution, then treated under three conditions: cold cream, SSD, and SSD combined with the tested protease. After 15 days of daily treatment, the mice were sacrificed and skin tissue samples were histopathologically examined using hematoxylin eosin, and Masson trichrome staining. RESULTS: The D10 protease hydrolyzed the proteinaceous components of eschars (fibrin, normal collagen, and denatured collagen) in vitro. Mice skins treated with protease and SSD mixture showed promising results, with more rapid healing than the other treatments. This group regenerated epidermis and dermis with newly formed granulated follicles, fibroblasts and blood capillaries in the dermis, and collagen fibers in the hypodermis. CONCLUSIONS: These results suggest that the serine protease produced by B. subtilis D10 promotes wound healing of mice skin burnt with HCl and restores the normal architectural pattern in a shorter time than the standard treatments.

Possible Protective Role of Whey Protein on the Rat's Liver Tissues Treated with Nandrolone decanoate.

BACKGROUND AND OBJECTIVE: Nandrolone and whey protein are used as supplementary food and athletic food. The aim of this study was to evaluate the possible histological and ultrastructural alterations in the liver of adult rats after treatment of the anabolic androgenic steroids (Nandrolone decanoate) and whey protein. MATERIALS AND METHODS: Twenty eight Wistar Albino male rats were used in the present study divided into 4 groups: Control group received 0.5 mL of saline solution by oral, Nandrolone group injected intramuscular (10 mg kg-1 b.wt./week for 3 months), whey protein group treated by oral (5 mg kg-1 b.wt./week for 3 months) and Nandrolone and whey protein group. At the end of the experimentation, all the rats were sacrificed and liver samples were processed for histological and ultrastructural examination. Haematoxylin and eosin stains for general histological examination and Mallory trichrome stain for collagen fibers. RESULTS: Light microscopy examination of the liver of the nandrolone group showed bleeding and widening of the blood sinusoids. Degeneration, vacuolation, coagulative necrosis and pyknotic nuclei were observed. In addition, increased collagen fibers were detected. Whey protein group showed more or less normal hepatocytes, blood sinusoids and collagen fibers. The nandrolone and whey protein group illustrated normal appearance of hepatocytes with vacuolation in some of the hepatocytes and normal blood sinusoids and collagen fibers were noticed. Electron microscopic examination of the nandrolone group showed depletion of the nuclear chromatin, damaged mitochondria, increased of lysosomes, some lipid droplets, damaged blood sinusoids and space of Disse and increased of Kupffer cells, whereas the whey protein group appeared normal. The nandrolone and whey protein group showed well developed hepatocytes, regular space of Disse and normal hepatic sinusoids. CONCLUSIONS: Whey protein may be ameliorate the hepatic architecture after treatment with nandrolone.