Human Gyrovirus Apoptin as a Potential Selective Anticancer Agent: An In Vitro Study

Background: Selective therapy has always been the main challenge in cancer treatments. Recently, it has been shown that Human Gyrovirus-derived protein apoptin (HGV-Apoptin) has selective cytotoxic effects on cancer cells similar to its homologue, Chicken Anemia Virus-derived Apoptin (CAV-Apoptin). However, apoptotic effects of Human Gyrovirus apoptin have been only evaluated on a few cancerous cell lines and need to be further investigated. In this study, we have evaluated the apoptotic effects of HGV-Apoptin and CAV-Apoptin expression on lung cancer (A549) and normal (HEK-293) cell lines, in order to provide more information about the specificity of these proteins on cancerous cells. Methods: Target cells were transfected by the calcium-phosphate precipitation method with constructed plasmids expressing HGV-Apoptin and CAV-Apoptin proteins as well as the control plasmid. Transfection efficiency was followed and imaged by fluorescence microscopy. Quantification of apoptosis was performed by flow cytometry. Measurements were compared by paired Student t-test. Results: Cells were successfully transfected with control and constructed plasmids. Flowcytometry analysis showed that A549 cells transfected with HGV-Apoptin and CAV-Apoptin expressing plasmids, undergone the apoptosis compared to A549 cells transfected with control plasmid (P<0.001). None of the plasmids could induce apoptosis in HEK-293 cells. Conclusion: Human Gyrovirus-derived apoptin (HGV-Apoptin) similar to its homologue, chicken anemia virus derived Apoptin (CAV-Apoptin) can induce apoptosis in Non-small-cell lung carcinoma cell line A549, but not in normal human embryonic kidney cell line HEK-293, which can be introduced as a promising novel specific antitumor agent

Overcoming Breast Cancer Drug Resistance: A Novel Approach Using siRNA-Mediated P-glycoprotein Downregulation to Enhance Vinorelbine Efficacy

Purpose: Cancer, the second leading cause of mortality worldwide, represents a global health challenge, primarily due to drug resistance. Vinorelbine is a chemotherapeutic agent that disrupts cancer cell growth by targeting microtubules and inducing apoptosis. However, drug resistance remains a formidable obstacle. This resistance is caused by various factors including genetic mutations, drug efflux mechanisms, and DNA repair systems. Resolution of this challenge requires an innovative approach. This study investigated the potential of small interfering RNA (siRNA) to target and downregulate a vinorelbine-resistant MCF-7/ADR breast cancer cell line. Methods: Cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) 10% fetal bovine serum/penicillin/streptomycin. An siRNA targeting ABCB1 was designed and synthesized, and the cells were transfected with siRNA at final concentrations of 10, 20, and 30 nM. The3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to assess cell viability. ABCB1 mRNA expression levels were determined by real-time polymerase chain reaction (PCR). Results: MCF-7 cells exhibited a higher sensitivity to vinorelbine than MCF-7/ADR cells. MCF-7/ADR cells exhibited resistance to vinorelbine at concentrations, 12.50 and 25.00 μM. Treatment with siRNA significantly reduced ABCB1 expression by 2.93-fold (P=0.0001). Similarly, co-treatment with siRNA and vinorelbine produced a substantial 2.89-fold decrease in ABCB1 gene expression in MCF-7 cells compared to that in MCF-7/ADR cells (P=0.0001). Conclusion: The results of the present study indicate that the concurrent use of siRNA and vinorelbine holds substantial promise as a therapeutic approach to overcome ABCB1-mediated multidrug resistance (MDR) in breast cancer. It is necessary to conduct comprehensive clinical trials to determine the true effectiveness of this combination therapy

Evaluation of Flow Cytometry and Kleihauer Techniques for Quantification of Fetomaternal Hemorrhage: A Prospective Cohort Study in Southwestern Iran

Background: Quantification of fetal red blood cells (RBCs) in maternal blood is of great importance to calculate appropriate dose of post-deliver anti D immunoglobulin in a rhesus D (RhD)-negative woman. Objective: The aim of this study is to evaluate a direct immunofluorescence flow cytometry technique in artificial and clinical samples and compared it to the Kleihauer-Betke test (KBT). Methods: This study was a prospective cohort design. Blood samples from 26 pregnant women who gave birth to RhD positive babies were tested using direct immunofluorescence flow cytometry and KBT techniques to determine the amount of FMH in the maternal circulation. The zone of D-positive cells was identified employing artificial samples including 0.3%, 0.6%, 1%, 1.5%, 2%, 5%, 10%, and 50% of D-positive fetal cells in D-negative maternal cells. Results: Analysis of 26 clinical samples for FMH showed consistent quantification with the flow cytometry and Kleihauer techniques. Although a good correlation was found between the KBT and flow cytometry results, in artificial samples containing more than 2% of fetal RhD positive cells, the flow cytometry results were closer to theoretical percentages. In a patient with FMH >4 mL, the FMH and consequently the required vial of Ig were overestimated using KBT. Conclusion: Most of the FMH calculated could have been neutralized by doses less than 625 IU, whereas the routine dose in Iran is more than double that amount (1500 IU). This achievement demonstrates that adjusting between the RhD immune globulin (RhDIg) dose and FMH size is inevitable

Unlocking Therapeutic Potential: Enhanced shRNA Delivery with Tat Peptide in the Human Respiratory Syncytial Virus Treatment

Purpose: This research investigated the development of short hairpin RNA (shRNA) molecules designed to target specific regions of the human respiratory syncytial virus (HRSV) M and F genes. The study aimed to assess the therapeutic potential of these shRNAs and evaluate the effectiveness of Tat peptide-mediated delivery in enhancing their functionality. Methods: We acquired isolates from pediatric patients experiencing respiratory illness then cultured in HEp-2 cells. We constructed plasmids expressing shRNAs. Tat peptide as a facilitator for shRNA plasmid delivery was used. The cytotoxicity of ribavirin, shRNA constructs, and control agents was assessed using the MTT assay. The transfection efficiency of Tat peptide-mediated shRNA delivery with that of lipofectamine 3000™ were compared. Finally, real-time PCR was employed to quantify HRSV replication in the treated cells. Results: Tat peptide-mediated delivery of shRNA plasmids significantly suppressed the expression of the M and F genes of HRSV compared to lipofectamine 3000™. This suppression was evident in both short-term experiments and scenarios involving stable shRNA expression. Furthermore, the combination of ribavirin with shRNA treatment resulted in a substantial reduction in viral load. Notably, the most pronounced antiviral effect was observed when both shRNAs were employed simultaneously. Conclusion: Our findings suggest that Tat peptide-mediated delivery of shRNA plasmids holds significant potential for achieving stable suppression of HRSV genes. This approach warrants further investigation as a potential gene therapy strategy for HRSV. By demonstrating promising results in vitro, this study highlights the need for future in vivo studies to comprehensively evaluate the therapeutic potential of this approach in a clinical setting

Emergence of Highly Multidrug-Resistant Bacteria Isolated from Patients with Infections Admitted to Public Hospitals in Southwest Iran

Background. The emergence of multidrug-resistant (MDR) microorganisms causing infections is increasing worldwide and becoming more serious in developing countries. Among those, Acinetobacter species are becoming prominent. Objectives. The aim of this study was to determine the rate of antimicrobial resistance of the bacteria causing infections, Acinetobacter species in particular, in local public hospitals in Firuzabad, Fars province, Iran. Methods. This cross-sectional study was performed on different clinical specimens collected from patients who were suspected of infections hospitalized from March 2016 to March 2019 in local hospitals of Firuzabad, Fars province, Iran. The bacterial isolates were identified following standard microbiological methods. Clinical and Laboratory Standards Institute guidelines were used to identify the antibiotic susceptibility of these isolates. Results. Overall, 1778 bacterial etiologies were isolated from 1533 patients diagnosed with infection. Of these, 1401 (78.8%) were Gram-negative and the remaining were Gram-positive bacteria. Escherichia coli (37.1%), Klebsiella spp. (13.9%), and Acinetobacter species (10.4%) were the most common isolated bacteria. Antibiotic sensitivity testing in this study showed a high resistance rate of Acinetobacter species to all antibiotics tested except Colistin. During the study period, the rate of infection with highly multidrug-resistant Acinetobacter species increased from 7.2% to 13.3%. Conclusions. This study highlights the emergence of MDR bacterial agents such as Acinetobacter species as a new threat in our region. However, a decrease in the rate of infection with Pseudomonas aeruginosa was noticeable.</jats:p

Emergence of Highly Multidrug-Resistant Bacteria Isolated from Patients with Infections Admitted to Public Hospitals in Southwest Iran

Background. The emergence of multidrug-resistant (MDR) microorganisms causing infections is increasing worldwide and becoming more serious in developing countries. Among those, Acinetobacter species are becoming prominent. Objectives. The aim of this study was to determine the rate of antimicrobial resistance of the bacteria causing infections, Acinetobacter species in particular, in local public hospitals in Firuzabad, Fars province, Iran. Methods. This cross-sectional study was performed on different clinical specimens collected from patients who were suspected of infections hospitalized from March 2016 to March 2019 in local hospitals of Firuzabad, Fars province, Iran. The bacterial isolates were identified following standard microbiological methods. Clinical and Laboratory Standards Institute guidelines were used to identify the antibiotic susceptibility of these isolates. Results. Overall, 1778 bacterial etiologies were isolated from 1533 patients diagnosed with infection. Of these, 1401 (78.8%) were Gram-negative and the remaining were Gram-positive bacteria. Escherichia coli (37.1%), Klebsiella spp. (13.9%), and Acinetobacter species (10.4%) were the most common isolated bacteria. Antibiotic sensitivity testing in this study showed a high resistance rate of Acinetobacter species to all antibiotics tested except Colistin. During the study period, the rate of infection with highly multidrug-resistant Acinetobacter species increased from 7.2% to 13.3%. Conclusions. This study highlights the emergence of MDR bacterial agents such as Acinetobacter species as a new threat in our region. However, a decrease in the rate of infection with Pseudomonas aeruginosa was noticeable