Iranian Biomedical Journal
Volume:26 Issue: 4, Jul 2022

Genetic factors are involved in the development, progression, and severity of non-alcoholic fatty liver disease (NAFLD). Polymorphisms in genes regulating liver functions may increase liver susceptibility to NAFLD. Therefore, we conducted this literature study to present recent findings on NAFLD-associated polymorphisms from published articles in PubMed from 2016 to 2021. From 69 selected research articles, 20 genes and 34 SNPs were reported to be associated with NAFLD. These mutated genes affect NAFLD by promoting liver steatosis (PNPLA3, MBOAT7, TM2SF6, PTPRD, FNDC5, IL-1B, PPARGC1A, UCP2, TCF7L2, SAMM50, IL-6, AGTR1, and NNMT), inflammation (PNPLA3, TNF-α, AGTR1, IL-17A, IL-1B, PTPRD, and GATAD2A), and fibrosis (IL-1B, PNPLA3, MBOAT7, TCF7L2, GATAD2A, IL-6, NNMT, UCP, AGTR1, and TM2SF6). The identification of these genetic factors helps to better understand the pathogenesis pathways of NAFLD.

Self-amplifying mRNA is the next-generation vaccine platform with the potential advantages in efficacy and speed of development against infectious diseases and cancer. The main aim was to present optimized and rapid methods for Semliki Forest virus (SFV)-PD self-amplifying mRNA (SAM) preparation, its packaging, and titer determination. These protocols are provided for producing and harvesting the high yields of virus replicon particle (VRP)-packaged SAM for vaccine studies.

pSFV-PD-EGFP plasmid was linearized and subjected to in vitro transcription. Different concentrations of SFV-PD SAM were first transfected into human embryonic kidney 293 cells (HEK-293) and baby hamster kidney cell line 21 (BHK-21) cell lines, and EGFP expression at different time points was evaluated by fluorescent microscopy. Replicon particle packaging was achieved by co-transfection of SFV-PD SAM and pSFV-Helper2 RNA into BHK-21 cells. The VRPs were concentrated using ultrafiltration with 100 kDa cut-off. The titers of replicon particles were determined by reverse transcription quantitative real-time PCR (RT-qPCR).

In vitro transcribed SAM encoding EGFP was successfully transfected and expressed in HEK-293 and BHK-21 cell lines. Higher levels of EGFP expression was observed in BHK-21 compared to HEK-293 cells showing more stable protein overexpression and VRP packaging. Using ultrafiltration, the high yields of purified SFV-PD-EGFP particles were rapidly obtained with only minor loss of replicon particles. Accurate and rapid titer determination of replication-deficient particles was achieved by RT-qPCR.

Using optimized methods for SAM transfection, VRP packaging, and concentration, high yields of SFV-PD VRPs could be produced and purified. The RT-qPCR demonstrated to be an accurate and rapid method for titer determination of replication deficient VRPs.

One of the most widely used anticancer agents is microbial L-ASNase. Herein, we assessed the biochemical and biological properties of an isolated L-ASNase from a Gram-negative bacteria strain, Escherichia coli MF-107.

Using garden asparagus, we obtained several bacterial isolates. These strains were further screened for L-ASNase activity. A promising bacterial isolate was selected for L-ASNase production and subsequent purification. The molecular weight of purified L-ASNase was determined.  The MTT assay was applied to assess the cytotoxic effect of the purified enzyme. Also, for caspase activity determination and the apoptotic effect of purified enzyme on in cells, we conducted a real-time PCR method.

The molecular weight of the enzyme was approximately 37 kDa. In the pH range of 7.5 to 8, the enzyme had considerable stability. At 35 °C, the purified L-ASNase optimum activity was recorded. The cytotoxic effect of the enzyme on treated cells was dose-dependent with an IC50 value of 5.7 IU/ml. The Bax gene expression considerably raised by 5.75-fold (p < 0.001) upon L-ASNase treatment. On the other hand, the anti-apoptotic Bcl-2 gene expression showed a 2.63-fold increase compared to the control (p < 0.05). It was detected that the mRNA levels of caspase-3 and p53 were considerably upregulated (5.93 and 1.85-fold, respectively). We did not find any alternation in the caspase-8 activity of the treated cells compared to untreated cells.

In this research, the proliferation of the breast cancer cells remarkably inhibited via the cytotoxic effect of isolated L-ASNase from microbial sources.

Recent years have brought notable progress in raising the efficiency of the reprogramming technique so that approaches have evolved from known transgenic factors to only a few miRNAs. Nevertheless, there is a poor understanding of both the key factors and biological networks underlying this reprogramming. The present study aimed to investigate the potential of miR-106b-5p in regulating spermatogonial stem cells (SSCs) to induced pluripotent stem cell (iPSC)-like cells.

We used SSCs because pluripotency is inducible in SSCs under defined culture conditions, and they have a few issues compared to other adult stem cells. As both signaling and post-transcriptional gene controls are critical for pluripotency regulation, we traced the expression of Oct-4, Sox-2, Klf-4, c-Myc, and Nanog (OSKMN). Besides, we considered miR-106b-5p targets using bioinformatic methods.

Our results showed that transfected SSCs with miR-106b-5p increased the expression of the OSKMN factors, which was significantly more than negative control groups. Moreover, using the functional miRNA enrichment analysis, online tools, and databases, we predicted that miR-106b-5p targeted a signaling pathway gene named MAPK1/ERK2, related to regulating stem cell pluripotency.

Together, our data suggest that miR-106b-5p regulates the reprogramming of SSCs into iPSC-like cells. Furthermore, noteworthy progress in the in vitro development of SSCs indicates promise reservoirs and opportunities for future clinical trials.

Glioblastoma is the most common primary malignant neoplasm of the central nervous system. Despite progress in diagnosis and treatment, glioblastoma still has a poor prognosis. This study aimed to examine whether a signature of three candidate miRNAs (i.e. hsa-let-7c-5p, hsa-miR-206-5p, and hsa-miR-1909-5p) can be used as a diagnostic biomarker for distinguishing glioblastoma from healthy brain tissues.

In this study, 50 formalin‐fixed paraffin‐embedded (FFPE) glioblastoma tissue samples and 50 healthy tissue samples adjacent to tumor were included. The expression of each candidate miRNA (i.e. hsa-let-7c-5p, hsa-miR-206-5p, and hsa-miR-1909-5p) was measured using quantitative reverse transcription PCR. To show the roles of each miRNA and their biological effects on glioblastoma development and clinicopathological characteristics, in silico tools were used. ROC curves were performed to assess the diagnostic accuracy of each miRNA.

Based on the results, hsa-let-7c-5p and hsa-miR-206-5p were downregulated, while hsa-miR-1909-5p was upregulated in glioblastoma tumors compared to healthy samples. No association was detected between the expression of each candidate miRNA and sex. Except for hsa-let-7c-5p, other miRNAs did not correlate with age status. ROC curve analysis indicated that the signature of candidate miRNAs is a potential biomarker distinguishing between glioblastoma and healthy samples. Only hsa-miR-206-5p suggested the association with poor prognosis in glioblastoma patients.

Our findings revealed that the signature of three miRNAs is capable of distinguishing glioblastoma tumor and healthy tissues. These results are beneficial for the clinical management of glioblastoma patients.

Autophagy induction has been shown to differ in magnitude depending on the mycobacterial species. However, few studies have investigated the specific autophagic capacity of different Mycobacterium tuberculosis (Mtb) strains in alveolar epithelial cells (ATs). This study aimed to elucidate the host autophagic response to different Mtb strains in ATs responsible for TB in the capital of Iran, Tehran.  

A549 cells were infected with three different Mtb clinical isolates (Beijing, NEW1, and CAS1/Delhi) and the reference strain H37Rv. Following RNA extraction, the expression of eight ATG genes, four mycobacterial genes, and three miRNAs was evaluated using quantitative RT-PCR.  

The results revealed that all four strains influenced the autophagy pathway in various ways at different magnitudes. The Beijing and H37Rv strains could inhibit autophagosome formation, whereas the CAS and NEW1 strains induced autophagosome formation. The expression of genes involved in the fusion of autophagosomes to lysosomes (LAMP1) indicated that all the studied strains impaired the autophagolysosomal fusion; this result is not unexpected as Mtb can block the autophagolysomal fusion. In addition, the Beijing and H37RV strains prevented the formation of autophagic vacuoles, besides mycobacterial targeting of lysosomes and protease activity.

This preliminary study improved our understanding of how Mtb manages to overcome the host immune system, such as autophagy, and evaluated the genes used by specific strains during this process. Further studies with a large number of Mtb strains, encompassing the other main Mtb lineages, are inevitable.

Enhancing the therapeutic profile of hydrophobic drugs using the development of biocompatible drug delivery systems is an urgent need. Many types of research have been conducted on graphene derivatives owing to their unique characteristics.  

In this survey, quercetin (QUER), a natural medicine, was loaded on carboxylated graphene oxide (GO), and cytotoxicity assay and the uptake of QUER into prostate cancer cells (PC3) were evaluated.

The release behavior of QUER was temperature- and pH-sensitive. Although QUER was loaded with high efficiency, the released rate was low (23.25% at pH 5.5 and 42 °C). The toxicity and intensity of fluorescence in the FREE QUER were higher than the loaded form.

High-capacity loading and controlled release of GO QUER can be recognized as a proper candidate in treating cancer.

Multiple sclerosis (MS) is the most prevalent neurological disability of young adults. Anti-inflammatory drugs have relative effects on MS. The anti-inflammatory and antioxidative effects of Zingiber officinale (ginger) have been proven in some experimental and clinical investigations. The aim of this study was to evaluate the effects of ginger extract on preventing myelin degradation in a rat model of MS.

Forty nine male Wistar rats were used in this study and divided into four control groups: the normal group, cuprizone-induced group, sham group (cuprizone [CPZ] + sodium carboxymethyl cellulose [NaCMC]), standard control group (fingolimod + cuprizone), including three experimental groups of CPZ, each receiving three different doses of ginger extract: 150, 300, and 600mg/kg /kg/day.

Ginger extract of 600 mg/kg prevented corpus callosum from demyelination; however, a significant difference was observed in the fingolimod group (p < 0.05). Difference in the CPZ group was quite significant (p < 0.05).

Treatment with ginger inhibited demyelination and alleviated remyelination of corpus callosum in rats. Therefore, it could serve as a therapeutic agent in the MS.