Stemness targeting of colorectal cell lines mediated by BAMLET and 5-Flourouracil.

PURPOSE: Stemness is the potential for self-renewal and repopulation causing the relapse, progression, and drug resistance of colorectal cancer. We investigated the effects of bovine alpha-lactalbumin made lethal to tumor cells and 5-Flourouracil consisting of bovine α-lactalbumin protein and oleic acid, on colorectal cancer cells on stemness. METHODS: The quantitative real-time polymerase chain reaction assessed the expression levels of stemness-related genes (c-myc, Lgr5, OCT4). Expression of stemness-related surface markers (CD44 and CD24) was also measured by the flow cytometry technique following the treatments. RESULTS: Our results indicated decreased expression levels of C-Myc, Lgr5, oct4 as the stemness-related genes (P < 0.0001), and reduced population of CD44+ as the stemness-related cell surface marker upon treatment with BAMLET and 5-Flourouracil. BAMLET inhibited the stemness more effectively than 5-Flourouracil (P < 0.0001). CONCLUSION: Based on the result, inhibition of the Stemness related-genes (C-Myc, Lgr5, Oct4) and the surface markers (CD 24+ and CD44+) is a promising therapeutic approach using BAMLET.

Epidemiological surveillance of respiratory viral infections in SARS-CoV-2-negative samples during COVID-19 pandemic in Iran.

BACKGROUND: To improve the patient care, public health surveillance, and infection control, it is crucial to identify the presence and frequency of the common respiratory infections in individuals with COVID-19 symptoms but tested negative for SARS-CoV-2. This study aimed to shed light on this during the COVID-19 pandemic in Iran. METHODS: In this cross-sectional study, a total of 1,002 patients with acute respiratory infection who had negative SARS-CoV-2 test results and referred to Valfajr Health Center, the National Collaborating Laboratory of Influenza and COVID-19 National Reference Laboratory at Pasteur Institute of Iran were recruited between January 2020 and January 2022. Nasopharyngeal and oropharyngeal swab samples were collected to detect 17 common respiratory viruses via TaqMan one-step real-time multiplex PCR. Demographic and clinical data of the participants were obtained from their electronic medical records. RESULTS: In total, 218 samples (21.8%) were tested positive for at least one respiratory virus infection. Most of the common investigated respiratory viruses belonged to the years 2020 and 2022. The number of investigated patients in 2021 was few, which highlights the impact of health measures following the COVID-19 pandemic in Iran. Influenza A was the most common virus (5.8%), while adenovirus had the lowest prevalence (0.1%). Although the rate of respiratory virus infection was higher in men (24%) compared to women (19.3%), this difference was not statistically significant (P = 0.069). The prevalence of respiratory viruses had an inverse association with increasing age, with the highest rate (55.6%) observed in the age group below 2 years and the lowest rate (12.7%) in those above 65 years. CONCLUSION: Our findings underscore the significance of adopting a comprehensive approach to respiratory infections detection and management. These results can be employed for the development of syndromic surveillance systems and implementation of the effective infection control measures. Furthermore, the results contribute to better understanding of the dynamics of respiratory viruses, both during pandemic periods and in non-pandemic contexts.

Evaluation of the antiviral potential of gemini surfactants against influenza virus H1N1.

Influenza A virus (IAV) affects human health worldwide as a high-risk disease. It can neither be easily controlled by current vaccines and nor be treated by conventional drugs. Gemini surfactants (GS) have shown several properties including antiviral activity. In this study, the antiviral capacity of some GS compounds with different levels of hydrophobicity was examined. The 50% cytotoxic (CC50) and non-cytotoxic (NCTC) concentrations of the compounds were determined by MTT method. The NCTCs, the same as effective concentrations (EC50s), were tested for the antiviral capacity against IAV in different combination treatments for 1 h incubation on MDCK cells. The HA and MTT assays were used to evaluate the virus titer and cell viabilities, respectively. The hemolytic activity of the compounds was also assessed using an HA inhibition assay. To evaluate the apoptotic effect of GS compounds, Annexin V-PI kit was used. The HA titers decreased between 1-6.5 logs, 1-4.5 logs, and 1-5.5 logs in simultaneous, pre- and post-penetration combination treatments, respectively. The cell viability values in all combination treatments were favorable. The HI assay indicated the hemolytic potential of GSs and their physical interaction with viral HA. The apoptosis test results highlighted anti-apoptotic capacity of the GS compounds alone and in the presence of influenza virus especially for the hydrophobic ones. Gemini surfactants were generally more efficacious in simultaneous treatment. Their antiviral potential may be attributed to their physical interaction with viral membrane or HA glycoprotein that disrupts viral particle or blocks viral entry to the cell and inhibits its propagation.

Antibacterial and Virucidal Evaluation of Ultrafine Wire Arc Sprayed German Silver Coatings.

Copper and its alloys are known as antimicrobial agents that can be used in public places; however, pure copper has a low wear resistance and tends to lose its gloss relatively fast and stainless steel is still more desirable because of its mechanical properties and stable appearance. In this research, German silver coatings, a copper-nickel alloy, are studied as a superior alternative for pure copper coatings. German silver coating on mild steel substrates and stainless steel with two different surface roughnesses was prepared and placed into water bath up to 6 months to investigate the corrosion and exposure effects on the antibacterial behavior. A range of techniques was used to study the microstructure, surface morphology and mechanical properties such as microhardness, coating bonding adhesion, surface roughness and wettability of the coating. Colony count method was used to measure the antibacterial properties, and samples were tested against influenza A virus to evaluate the virucidal activity. The coating thickness was around 130 µm and contained 15% pores and oxides with splats forming inside the coating structure. Inside each splat, columnar grains could be seen with an average of 700 nm width and 4 µm length. The bonding strength of the coating was about 15 MPa, the hardness of coatings was about 180 HV, and the average surface roughness of the as-sprayed samples was about 10 µm. German silver coatings can destroy both Staphylococcus aureus and Escherichia coli by more than 90% after 6 h of exposure time, and it also has a high-level of virucidal activity against influenza A virus after 2 h exposure time. Antibacterial behavior did not show any significant changes after 6 months of immersing samples in water bath. Thus, thermally sprayed German silver coatings exhibited silvery color for a long period of time, while its antimicrobial efficiency was comparable to pure copper coatings. Supplementary Information: The online version contains supplementary material available at 10.1007/s11666-022-01528-4.

In vitro anti-influenza assessment of anionic compounds ascorbate, acetate and citrate.

BACKGROUND: Influenza A virus (IAV) infection remains a serious public health threat. Due to drug resistance and side effects of the conventional antiviral drugs, repurposing the available natural compounds with high tolerability and fewer side effects has attracted researchers' attention. The aim of this study was to screen in vitro anti-influenza activity of three anionic compounds ascorbate, acetate, and citrate. METHODS: The non-cytotoxic concentration of the compounds was determined by MTT assay and examined for the activity against IAV in simultaneous, pre-, and post-penetration combination treatments over 1 h incubation on Madin-Darby Canine Kidney (MDCK) cell line. The virus titer and viral load were determined using hemagglutination assay (HA) and qPCR, respectively. Few pro-inflammatory and anti-inflammatory cytokines were evaluated at RNA and protein levels by qPCR and ELISA, respectively. RESULTS: The non-cytotoxic concentrations of the ascorbate (200 mg/ml), acetate and citrate (both 3 mg/ml) reduced the viral titer by 6.5, 4.5, and 1.5 logs in the simultaneous combination treatment. The M protein gene copy number decreased significantly in simultaneous treatment (P < 0.01). The expression of cytokines was also affected by the treatment of these compounds. CONCLUSIONS: These anionic compounds could affect the influenza virus load, thereby reducing pro-inflammatory cytokines and increasing anti-inflammatory cytokines levels.

Recent Advances in the Control of Clinically Important Biofilms.

Biofilms are complex structures formed by bacteria, fungi, or even viruses on biotic and abiotic surfaces, and they can be found in almost any part of the human body. The prevalence of biofilm-associated diseases has increased in recent years, mainly because of the frequent use of indwelling medical devices that create opportunities for clinically important bacteria and fungi to form biofilms either on the device or on the neighboring tissues. As a result of their resistance to antibiotics and host immunity factors, biofilms have been associated with the development or persistence of several clinically important diseases. The inability to completely eradicate biofilms drastically increases the burden of disease on both the patient and the healthcare system. Therefore, it is crucial to develop innovative ways to tackle the growth and development of biofilms. This review focuses on dental- and implant-associated biofilm infections, their prevalence in humans, and potential therapeutic intervention strategies, including the recent advances in pharmacology and biomedical engineering. It lists current strategies used to control the formation of clinically important biofilms, including novel antibiotics and their carriers, antiseptics and disinfectants, small molecule anti-biofilm agents, surface treatment strategies, and nanostructure functionalization, as well as multifunctional coatings particularly suitable for providing antibacterial effects to the surface of implants, to treat either dental- or implant-related bacterial infections.

Association of the host genetic factors, hypercholesterolemia and diabetes with mild influenza in an Iranian population.

BACKGROUND: Variation in host genetic factors may result in variation in the host immune response to the infection. Some chronic diseases may also affect individuals' susceptibility to infectious diseases. The aim of this study was to evaluate the association of the host genetic factors mostly involved in inflammation, as well as hypercholesterolemia and diabetes with mild flu in an Iranian population. METHODS: In this cross-sectional study, nasopharyngeal swab samples were collected from 93 patients referred to primary care centers of Markazi, Semnan, and Zanjan provinces (central Iran) due to flu-like symptoms between March 2015 and December 2018. Of these, PCR test identified 49 influenza A/H1N1 and 44 flu-negative individuals. Twelve single-nucleotide polymorphisms (SNPs) in RPAIN, FCGR2A, MBL-2, CD55, C1QBP, IL-10, TNF-α and an unknown gene were genotyped using iPLEX GOLD SNP genotyping analysis. Hypercholesterolemia and diabetes status was determined based on the physician diagnosis. Association of the host genetic variants, hypercholesterolemia and diabetes with mild A/H1N1 flu was assessed with univariable and multivariable logistic regression analysis as implemented in Stata software (v.14). Statistical tests were considered as significant at 0.05 levels. RESULTS: Frequency of diabetes and hypercholesterolemia, as well as participants mean age was significantly higher in the flu-negative rather than the flu-positive group. Of 12 SNPs, nine did not show any significant association with mild flu in our study (rs1801274, rs1800451, rs2564978, rs361525, rs1800450, rs1800871, rs1800872, rs1800896, rs1800629). Possessing G vs. A allele in two SNPs (rs3786054 and rs8070740) was associated with a threefold increase in the chance of mild flu when compared to flu-negative patients (95% CI: 1.1, 22.0). Possessing C allele (vs. A) in the rs9856661 locus also increased the chance of mild flu up to 2 folds (95% CI: 1.0, 10.0). CONCLUSION: The results showed that possessing the G allele in either rs3786054 or rs8070740 loci in C1QBP and RPAIN genes, respectively, increased the risk of H1N1 infection up to 3.3 folds, regardless of the patient's age, BMI, diabetes, and hypercholesterolemia. Complementary functional genomic studies would shed more light on the underlying mechanism of human immunity associated with these genetic markers. The identified genetic factors may have the same role in susceptibility to similar respiratory infections with RNA viruses, like SARS, MERS and COVID-19. Future genetic association studies targeting these RNA viruses, especially COVID-19 is recommended. Studies on other ethnic groups would also shed light on possible ethnic variations in genetic susceptibility to respiratory RNA viruses. Trial registry IR.PII.REC.1399.063.

Autophagy, Unfolded Protein Response, and Neuropilin-1 Cross-Talk in SARS-CoV-2 Infection: What Can Be Learned from Other Coronaviruses.

The COVID-19 pandemic is caused by the 2019-nCoV/SARS-CoV-2 virus. This severe acute respiratory syndrome is currently a global health emergency and needs much effort to generate an urgent practical treatment to reduce COVID-19 complications and mortality in humans. Viral infection activates various cellular responses in infected cells, including cellular stress responses such as unfolded protein response (UPR) and autophagy, following the inhibition of mTOR. Both UPR and autophagy mechanisms are involved in cellular and tissue homeostasis, apoptosis, innate immunity modulation, and clearance of pathogens such as viral particles. However, during an evolutionary arms race, viruses gain the ability to subvert autophagy and UPR for their benefit. SARS-CoV-2 can enter host cells through binding to cell surface receptors, including angiotensin-converting enzyme 2 (ACE2) and neuropilin-1 (NRP1). ACE2 blockage increases autophagy through mTOR inhibition, leading to gastrointestinal complications during SARS-CoV-2 virus infection. NRP1 is also regulated by the mTOR pathway. An increased NRP1 can enhance the susceptibility of immune system dendritic cells (DCs) to SARS-CoV-2 and induce cytokine storm, which is related to high COVID-19 mortality. Therefore, signaling pathways such as mTOR, UPR, and autophagy may be potential therapeutic targets for COVID-19. Hence, extensive investigations are required to confirm these potentials. Since there is currently no specific treatment for COVID-19 infection, we sought to review and discuss the important roles of autophagy, UPR, and mTOR mechanisms in the regulation of cellular responses to coronavirus infection to help identify new antiviral modalities against SARS-CoV-2 virus.

Emerging Advances of Nanotechnology in Drug and Vaccine Delivery against Viral Associated Respiratory Infectious Diseases (VARID).

Viral-associated respiratory infectious diseases are one of the most prominent subsets of respiratory failures, known as viral respiratory infections (VRI). VRIs are proceeded by an infection caused by viruses infecting the respiratory system. For the past 100 years, viral associated respiratory epidemics have been the most common cause of infectious disease worldwide. Due to several drawbacks of the current anti-viral treatments, such as drug resistance generation and non-targeting of viral proteins, the development of novel nanotherapeutic or nano-vaccine strategies can be considered essential. Due to their specific physical and biological properties, nanoparticles hold promising opportunities for both anti-viral treatments and vaccines against viral infections. Besides the specific physiological properties of the respiratory system, there is a significant demand for utilizing nano-designs in the production of vaccines or antiviral agents for airway-localized administration. SARS-CoV-2, as an immediate example of respiratory viruses, is an enveloped, positive-sense, single-stranded RNA virus belonging to the coronaviridae family. COVID-19 can lead to acute respiratory distress syndrome, similarly to other members of the coronaviridae. Hence, reviewing the current and past emerging nanotechnology-based medications on similar respiratory viral diseases can identify pathways towards generating novel SARS-CoV-2 nanotherapeutics and/or nano-vaccines.

Oncolytic effects of Hitchner B1 strain of newcastle disease virus against cervical cancer cell proliferation is mediated by the increased expression of cytochrome C, autophagy and apoptotic pathways.

Newcastle disease virus (NDV) is a potential oncolytic virus for the cancer treatment due to its ability to replicate in tumor cells. The aim of this study was to evaluate the in vitro anticancer properties of Hitchner B1 (HB1) strain of NDV on TC-1 cell line and underlying molecular mechanisms. The cytolytic effects of oncolytic HB1 strain of NDV was determined by lactate dehydrogenase (LDH) release assay. Apoptosis, intracellular reactive oxygen species (ROS) levels, cleaved caspase-3 and autophagy were evaluated by flow cytometry. Cytochrome-C and survivin protein levels were distinguished by Enzyme-Linked Immunosorbent Assay (ELISA). Our results from LDH method showed that the viability of the TC-1 cell line following HB1 NDV infection was dose-dependent and decreased significantly with increasing the dose of HB1 NDV infection (MOIs: 5, 10, and 15). Other evaluations also revealed that HB1 strain of NDV potentially led to the ROS production, and apoptosis and autophagy induction in TC-1 cell line in a dose-dependent manner. The in vitro experiments also presented that NDV treatment significantly up-regulated the expression of cytochrome-C and down-regulated the expression of survivin, as detected by ELISA assay. Our results confirmed that the HB1 NDV could be introduced as a powerful candidate for the therapy of cervical cancer. However, further examinations are needed to explain the underlying mechanisms of the HB1 NDV against TC-1 cell line and cervical cancer.

Association of polymorphisms in inflammatory cytokines encoding genes with severe cases of influenza A/H1N1 and B in an Iranian population.

BACKGROUND: The increased levels of blood cytokines is the main immunopathological process that were attributed to severe clinical outcomes in cases of influenza A, influenza B and people with influenza-like illness (ILI). Functional genetic polymorphisms caused by single nucleotide polymorphisms (SNPs) in inflammatory cytokines genes can influence their functions either qualitatively or quantitatively, which is associated with the possibility of severe influenza infections. The aim of the present case-control study was to investigate the association of polymorphisms in inflammatory cytokines genes with influenza patients and ILI group in an Iranian population. METHODS: Total number of 30 influenza B, 50 influenza A (H1N1) and 96 ILI inpatient individuals were confirmed by Real-time RT-PCR and HI assays. The genotype determination was assessed for defined SNPs in IL-1ß, IL-17, IL-10 and IL-28 genes. RESULTS: The frequencies of the IL-1ß rs16944 (P = 0.007) and IL-17 rs2275913 (P = 0.006) genotypes were associated with severe influenza disease, while the frequencies of IL-10 rs1800872 and IL-28 rs8099917 were not associated with the disease (P > 0.05). Also, the absence of A allele in IL-17 rs2275913 SNP increased the risk of influenza A (H1N1) infection (P = 0.008). CONCLUSIONS: This study demonstrated that influenza A- (H1N1) and B-infected patients and also ILI controls have different profiles of immune parameters, and individuals carrying the specific cytokine-derived polymorphisms may show different immune responses towards severe outcome.

Autophagy: The multi-purpose bridge in viral infections and host cells.

Autophagy signaling pathway is involved in cellular homeostasis, developmental processes, cellular stress responses, and immune pathways. The aim of this review is to summarize the relationship between autophagy and viruses. It is not possible to be fully comprehensive, or to provide a complete "overview of all viruses". In this review, we will focus on the interaction of autophagy and viruses and survey how human viruses exploit multiple steps in the autophagy pathway to help viral propagation and escape immune response. We discuss the role that macroautophagy plays in cells infected with hepatitis C virus, hepatitis B virus, rotavirus gastroenteritis, immune cells infected with human immunodeficiency virus, and viral respiratory tract infections both influenza virus and coronavirus.

Experimental validation and computational modeling of anti-influenza effects of quercetin-3-O-α-L-rhamnopyranoside from indigenous south African medicinal plant Rapanea melanophloeos.

BACKGROUND: Influenza A virus (IAV) is still a major health threat. The clinical manifestations of this infection are related to immune dysregulation, which causes morbidity and mortality. The usage of traditional medication with immunomodulatory properties against influenza infection has been increased recently. Our previous study showed antiviral activity of quercetin-3-O-α-L-rhamnopyranoside (Q3R) isolated from Rapanea melanophloeos (RM) (L.) Mez (family Myrsinaceae) against H1N1 (A/PR/8/34) infection. This study aimed to confirm the wider range of immunomodulatory effect of Q3R on selective pro- and anti-inflammatory cytokines against IAV in vitro, to evaluate the effect of Q3R on apoptosis pathway in combination with H1N1, also to assess the physical interaction of Q3R with virus glycoproteins and RhoA protein using computational docking. METHODS: MDCK cells were exposed to Q3R and 100CCID50/100 µl of H1N1 in combined treatments (co-, pre- and post-penetration treatments). The treatments were tested for the cytokines evaluation at RNA and protein levels by qPCR and ELISA, respectively. In another set of treatment, apoptosis was examined by detecting RhoA GTPase protein and caspase-3 activity. Molecular docking was used as a tool for evaluation of the potential anti-influenza activity of Q3R. RESULTS: The expressions of cytokines in both genome and protein levels were significantly affected by Q3R treatment. It was shown that Q3R was much more effective against influenza when it was applied in co-penetration treatment. Q3R in combination with H1N1 increased caspase-3 activity while decreasing RhoA activation. The molecular docking results showed strong binding ability of Q3R with M2 transmembrane, Neuraminidase of 2009 pandemic H1N1, N1 and H1 of PR/8/1934 and Human RhoA proteins, with docking energy of - 10.81, - 10.47, - 9.52, - 9.24 and - 8.78 Kcal/mol, respectively. CONCLUSIONS: Quercetin-3-O-α-L-rhamnopyranoside from RM was significantly effective against influenza infection by immunomodulatory properties, affecting the apoptosis pathway and binding ability to viral receptors M2 transmembrane and Neuraminidase of 2009 pandemic H1N1 and human RhoA cellular protein. Further research will focus on detecting the detailed specific mechanism of Q3R in virus-host interactions.

Evaluation of autophagy induction on HEV 239 vaccine immune response in a mouse model.

Hepatitis E virus (HEV) infection remains a serious threat to life and productivity in developing world. Vaccine seems to be an effective, safe, and affordable approach to address HEV disease burden. The HEV genome consists of three open reading frames (ORFs). Of these, ORF2 encodes a single structural protein (pORF2) for the HEV capsid which has been studied extensively as vaccine candidates. Recently, it has been recognized that autophagy plays an important role in innate and adaptive immunity defense against intracellular pathogens. This mechanism could therefore promote a protective immune response by inducing CD4+ and CD8+ T cells. In this study, HEV 239 and Beclin1 proteins were expressed in prokaryotic host cell [Escherichia coli (BL21)]. HEV 239 protein with different formulations (+Alum, +Beclin1, and +Alum-Beclin1) were used as candidate vaccines and administrated subcutaneously in BALB/c mice on 0, 14, and 28 days. Finally, elicited cellular and humoral immunity were evaluated. Taken together, although our results indicated that mice immunized with HEV 239 protein formulated with Alum, Beclin1, and Alum + Beclin1 displayed humoral and cellular response that was not significant in comparison with each other (P > 0.05); whereas they were significant while compared with control groups (P < 0.05). A comprehensive understanding of the intricate interplay between autophagy and immune response remains to be unraveled. Further study will clear the detailed impact of autophagy manipulation to enhance vaccine efficacy and boost the immune responses against the disease. © 2018 IUBMB Life, 70(3):207-214, 2018.

Immunomodulatory properties of quercetin-3-O-α-L-rhamnopyranoside from Rapanea melanophloeos against influenza a virus.

BACKGROUND: Influenza infection is a major public health threat. The role of influenza A virus-induced inflammatory response in severe cases of this disease is widely recognized. Drug resistance and side effects of chemical treatments have been observed, resulting in increased interest in alternative use of herbal medications for prophylaxis against this infection. The South African medicinal plant, Rapanea melanophloeos (RM) (L.) Mez of the family Myrsinaceae was selected owing to its traditional use for the treatment of several diseases such as respiratory ailments and also previous preliminary studies of anti-influenza activity of its methanolic extract. The aim of this study was to investigate the immunomodulatory properties of a glycoside flavone isolated from RM against influenza A virus. METHODS: The non-cytotoxic concentration of the quercetin-3-O-α-L-rhamnopyranoside (Q3R) was determined by MTT assay and tested for activity against influenza A virus (IAV) in simultaneous, pre-penetration and post-penetration combination treatments over 1 h incubation on MDCK cells. The virus titer and viral load targeting NP and M2 viral genes were determined using HA and qPCR, respectively. TNF-α and IL-27 as pro- and anti-inflammatory cytokines were measured at RNA and protein levels by qPCR and ELISA, respectively. RESULTS: Quercetin-3-O-α-L-rhamnopyranoside at 150 µg/ml decreased the viral titer by 6 logs (p < 0.01) in the simultaneous procedure. The NP and M2 genes copy numbers as viral target genes, calculated based on the Ct values and standard formula, significantly decreased in simultaneous treatment (p < 0.01). The expression of cytokines was also considerably affected by the compound treatment. CONCLUSIONS: This is the first report of quercetin-3-O-α-L-rhamnopyranoside from RM and its immunomodulatory properties against influenza A virus. Further research will focus on detecting the specific mechanism of virus-host interactions.

South African medicinal plant extracts active against influenza A virus.

BACKGROUND: Influenza infection remains a major health threat for animals and humans which crucially requires effective antiviral remedies. The usage of herbal medications as readily available alternatives for their compatibility with the body and fewer side effects compared to synthetic chemical treatments has become popular globally. The aim of this study was to investigate and screen in vitro anti-influenza activity of extracts of five South African medicinal plants, namely Tabernaemontana ventricosa, Cussonia spicata, Rapanea melanophloeos, Pittosporum viridiflorum and Clerodendrum glabrum, species which are used traditionally for the treatment of several diseases such as inflammatory and respiratory diseases. METHODS: Methanol, ethanol (100% and 30%), acetone, hot and cold water extracts of the powdered plants leaves were obtained by standard methods. The cytotoxicity was determined by the MTT colorimetric assay on MDCK cells. The concentrations below CC50 values were tested for antiviral activity against influenza A virus (IAV) in different combination treatments. The effect of extracts on viral surface glycoproteins and viral titer were tested by HI and HA virological assays, respectively. RESULTS: Based on the applied methods, the most effective results against IAV were obtained from Rapanea melanophloeos methanol leaf extract (EC50 = 113.3 µg/ml) and Pittosporum viridiflorum methanol, 100% and 30% ethanol and acetone leaf extracts (EC50 values = 3.6, 3.4, 19.2, 82.3 µg/ml, respectively) in all types of combined treatments especially in pre- and post-penetration combined treatments with highly significant effects against viral titer (P ≤ 0.01). CONCLUSION: The outcomes offer for the first time a scientific basis for the use of extracts of Rapanea melanophloeos and Pittosporum viridiflorum against IAV. It is worth focusing on the isolation and identification of effective active compounds and elucidating the mechanism of action from these species. However, Tabernaemontana ventricosa, Cussonia spicata and Clerodendrum glabrum leaf extracts were ineffective in vitro in this study.

Association of IFITM3 rs12252 polymorphisms, BMI, diabetes, and hypercholesterolemia with mild flu in an Iranian population.

BACKGROUND: IFITM3 has been suggested to be associated with infection in some ethnic groups. Diabetes and hypercholesterolemia are also important clinical conditions that can predispose individuals to infection. The aim of this study was to investigate the association of rs12252 C polymorphism, BMI, diabetes, and hypercholesterolemia with mild flu in an Iranian population. METHODS: We conducted a case-control study, including 79 mild flu and 125 flu-negative individuals attending primary care centers of three provinces of Iran (i.e, Markazi, Semnan, and Zanjan). Pharyngeal swab specimens were collected from all participants, and were subjected to RNA and DNA extractions for Real-time PCR and PCR tests. All PCR products were then sequenced to find T/C polymorphisms in the rs12252 region. Data on demographic, anthropometric, and clinical variables were collected from participants' medical records available in the primary care centers. The data was analyzed using DNASIS (v. 2.5) and Stata (v.11) software. RESULTS: All participants were of Fars ethnic background. The allele frequency for rs12252-C was found to be 9.49% among cases and 2.40% among controls. Carriers of the rs12252 C allele (CT + CC genotypes) showed 5.92 folds increase in the risk of mild flu comparing to the T allele homozygotes (P value: 0.007). We also found a significant positive association between rs12252 C allele heterozygote and mild flu (OR: 7.62, P value: 0.008), but not in C allele homozygote group (OR: 2.71, P value: 0.406). Similarly, we did not find a significant association between mild flu and BMI (OR: 1.06, P value: 0.087), diabetes (OR: 0.61, P value: 0.392), and hypercholesterolemia (OR: 0.50, P value: 0.393) in multivariable logistic regression. CONCLUSIONS: This is the first study evaluating the association between rs12252 polymorphisms, diabetes, hypercholesterolemia, and BMI and susceptibility to mild flu in an Iranian population. Our results suggest a significant positive association between mild flu and rs12252 C allele heterozygous and carriage. Future replication of the strong association observed here between rs12252 C allele carriage and mild flu might candidate this polymorphism as a genetic marker for early screening of susceptibility to mild flu. Lack of significant association between C allele homozygous and mild flu, observed in this study, might be the result of small sample size in this group. TRIAL REGISTRATION: IR.PII.REC.1395.3.

Edible bird's nest modulate intracellular molecular pathways of influenza A virus infected cells.

BACKGROUND: Edible Bird's Nest (EBN) as a popular traditional Chinese medicine is believed to have health enhancing and antiviral activities against influenza A virus (IAV); however, the molecular mechanism behind therapeutic effects of EBN is not well characterized. METHODS: In this study, EBNs that underwent different enzymatic preparation were tested against IAV infected cells. 50% cytotoxic concentration (CC50) and 50% inhibitory concentration (IC50) of the EBNs against IAV strain A/Puerto Rico/8/1934(H1N1) were determined by HA and MTT assays. Subsequently, the sialic acid content of the used EBNs were analyzed by fluorometric HPLC. Western Blotting and immunofluorescent staining were used to investigate the effects of EBNs on early endosomal trafficking and autophagy process of influenza virus. RESULTS: This study showed that post inoculations of EBNs after enzymatic preparations have the highest efficacy to inhibit IAV. While CC50 of the tested EBNs ranged from 27.5-32 mg/ml, the IC50 of these compounds ranged between 2.5-4.9 mg/ml. EBNs could inhibit IAV as efficient as commercial antiviral agents, such as amantadine and oseltamivir with different mechanisms of action against IAV. The antiviral activity of these EBNs correlated with the content of N-acetyl neuraminic acid. EBNs could affect early endosomal trafficking of the virus by reducing Rab5 and RhoA GTPase proteins and also reoriented actin cytoskeleton of IAV infected cells. In addition, for the first time this study showed that EBNs can inhibit intracellular autophagy process of IAV life cycle as evidenced by reduction of LC3-II and increasing of lysosomal degradation. CONCLUSIONS: The results procured in this study support the potential of EBNs as supplementary medication or alternative to antiviral agents to inhibit influenza infections. Evidently, EBNs can be a promising antiviral agent; however, these natural compounds should be screened for their metabolites prior to usage as therapeutic approach.

Comparison between MDCK and MDCK-SIAT1 cell lines as preferred host for cell culture-based influenza vaccine production.

OBJECTIVES: To evaluate MDCK and MDCK-SIAT1 cell lines for their ability to produce the yield of influenza virus in different Multiplicities of Infection. RESULTS: Yields obtained for influenza virus H1N1 grown in MDCK-SIAT1 cell was almost the same as MDCK; however, H3N2 virus grown in MDCK-SIAT1 had lower viral titers in comparison with MDCK cells. The optimized MOIs to infect the cells on plates and microcarrier were selected 0.01 and 0.1 for H1N1 and 0.001 and 0.01 for H3N2, respectively. CONCLUSIONS: MDCK-SIAT1 cells may be considered as an alternative mean to manufacture cell-based flu vaccine, especially for the human strains (H1N1), due to its antigenic stability and high titer of influenza virus production.