Silencing E6/E7 Oncoproteins in SiHa Cells Treated with siRNAs and Oroxylum indicum Extracts Induced Apoptosis by Upregulating p53/pRb Pathways.

E6 and E7 human papillomavirus (HPV) oncoproteins play a significant role in the malignant transformation of infected cervical cancer cells via suppression of tumour suppressor pathways by targeting p53 and pRb, respectively. This study aimed to investigate the anticancer effects of Oroxylum indicum (OI) leaves' methanol extract on SiHa cervical cancer cells. Expression of apoptosis-related proteins (Bcl-2, caspase (cas)-3, and cas-9), viral oncoproteins (E6 and E7), and tumour suppressor proteins (p53 and pRb) were evaluated using western blot analysis before and after E6/E7 small interfering RNAs (siRNAs) transfection. In addition, the E6/E7 mRNA expression levels were assessed with real-time (RT)-PCR. The present study showed that the OI extract effectively hindered the proliferation of SiHa cells and instigated increments of cas-3 and cas-9 expressions but decreased the Bcl-2 expressions. The OI extract inhibited E6/E7 viral oncoproteins, leading to upregulation of p53 and pRb tumour suppressor genes in SiHa cells. Additionally, combinatorial treatment of OI extract and gossypin flavonoid induced restorations of p53 and pRb. Treatment with OI extract in siRNA-transfected cells also further suppressed E6/E7 expression levels and further upregulations of p53 and pRb proteins. In conclusion, OI extract treatment on siRNAs-transfected SiHa cells can additively and effectively block E6- and E7-dependent p53 and pRb degradations. All these data suggest that OI could be explored for its chemotherapeutic potential in cervical cancer cells with HPV-integrated genomes.

Distribution of 22 Single Nucleotide Polymorphisms in 13 Cytokine Genes in Malays, Chinese, and Indians in Peninsular Malaysia.

Introduction: Cytokines are cell signaling glycoproteins that are particularly important in immunity and inflammatory responses. Therefore, variations, such as single nucleotide polymorphisms (SNPs), in genes encoding for cytokines may have important consequences for their roles in health. Materials and Methods: A total of 222 unrelated, healthy, and un-admixed Malays (n = 97), Chinese (n = 77), and Indians (n = 48) with a median age of 30 years old (range 21-50) were typed for 22 cytokine gene SNPs: IL-1α -889 T/C, IL-1ß (-511 T/C, +3962 T/C), IL-1R pst1 1970 T/C, IL-1RA mspa1 11100 T/C, IL-4Rα +1902 G/A, IL-12 - 1188 C/A, IFN-γ +874 A/T, TGF-ß (cdn 10 C/T, cdn 25 G/C), TNF-α (-308 A/G, -238 A/G) IL-2 (+166 G/T, -330 T/G), IL-4 (-1098 T/G, -590 T/C, -33 T/C), IL-6 (-174 C/G, nt565 G/A), and IL-10 (-1082 G/A, -819 C/T, -592 A/C). This involved using well-established polymerase chain reaction procedures with sequence-specific primers and restriction fragment length polymorphism methods. Results: The majority of the screened cytokine gene SNPs are polymorphic in all three ethnicities. Exceptions include TGF-ß cdn 25 (G/C), IL-1ß +3962 (T/C), and TNF-α -238 (A/G), which were all observed to be monomorphic in Malays, Chinese and Indians. Many of the analyzed cytokine gene SNP genotypes deviated from Hardy-Weinberg equilibrium and the three ethnic study groups were all well-separated from reference Asian, African and European populations in a principal component analysis plot. Conclusion: We successfully typed 22 SNPs in 13 cytokine genes from genetic material collected from unrelated and un-admixed Malay, Chinese and Indian individuals in Peninsular Malaysia. These new cytokine gene population datasets reveal interesting contrasts with other populations.

Bioactivity and Pharmacological Potential of Date Palm (Phoenix dactylifera L.) Against Pandemic COVID-19: a Comprehensive Review.

A novel coronavirus disease (COVID-19) or severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2), transmitted from person to person, has quickly emerged as the pandemic responsible for the current global health crisis. This infection has been declared a global pandemic, resulting in a concerning number of deaths as well as complications post-infection, primarily among vulnerable groups particularly older people and those with multiple comorbidities. In this article, we review the most recent research on the role of date palm (Phoenix dactylifera L.) fruits (DPFs) to prevent or treat COVID-19 infection. The mechanisms underlying this preventive or therapeutic effect are also discussed in terms of bioactivity potentials in date palm, e.g., antimicrobial, antioxidant, anticancer, anti-diabetic, anti-inflammatory, neuroprotective, and hemolytic potential, as well as prospect against COVID-19 disease and the potential product development. Therefore, it can be concluded that regular consumption of DPFs may be associated with a lower risk of some chronic diseases. Indeed, DPFs have been widely used in folk medicine since ancient times to treat a variety of health conditions, demonstrating the importance of DPFs as a nutraceutical and source of functional nourishment. This comprehensive review aims to summarize the majority of the research on DPFs in terms of nutrient content and biologically active components such as phenolic compounds, with an emphasis on their roles in improving overall health as well as the potential product development to ensure consumers' satisfaction in a current pandemic situation. In conclusion, DPFs can be given to COVID-19 patients as a safe and effective add-on medication or supplement in addition to routine treatments.

Synergetic Influence of Bismuth Oxide Nanoparticles, Cisplatin and Baicalein-Rich Fraction on Reactive Oxygen Species Generation and Radiosensitization Effects for Clinical Radiotherapy Beams.

PURPOSE: This study aimed to quantify synergetic effects induced by bismuth oxide nanoparticles (BiONPs), cisplatin (Cis) and baicalein-rich fraction (BRF) natural-based agent on the reactive oxygen species (ROS) generation and radiosensitization effects under irradiation of clinical radiotherapy beams of photon, electron and HDR-brachytherapy. The combined therapeutic responses of each compound and clinical radiotherapy beam were evaluated on breast cancer and normal fibroblast cell line. METHODS: In this study, individual BiONPs, Cis, and BRF, as well as combinations of BiONPs-Cis (BC), BiONPs-BRF (BB) and BiONPs-Cis-BRF (BCB) were treated to the cells before irradiation using HDR brachytherapy with 0.38 MeV iridium-192 source, 6 MV photon beam and 6 MeV electron beam. The individual or synergetic effects from the application of the treatment components during the radiotherapy were elucidated by quantifying the ROS generation and radiosensitization effects on MCF-7 and MDA-MB-231 breast cancer cell lines as well as NIH/3T3 normal cell line. RESULTS: The ROS generated in the presence of Cis stimulated the most substantial amount of ROS compared to the BiONPs and BRF. Meanwhile, the combination of the components had induced the higher ROS levels for photon beam than the brachytherapy and electron beam. The highest ROS enhancement relative to the control is attributable to the presence of BC combination in MDA-MB-231 cells, in comparison to the BB and BCB combinations. The radiosensitization effects which were quantified using the sensitization enhancement ratio (SER) indicate the highest value by BC in MCF-7 cells, followed by BCB and BB treatment. The radiosensitization effects are found to be more prominent for brachytherapy in comparison to photon and electron beam. CONCLUSION: The BiONPs, Cis and BRF are the potential radiosensitizers that could improve the efficiency of radiotherapy to eradicate the cancer cells. The combination of these potent radiosensitizers might produce multiple effects when applied in radiotherapy. The BC combination is found to have the highest SER, followed by the BCB combination. This study is also the first to investigate the effect of BRF in combination with BiONPs (BB) and BC (BCB) treatments.

Radiosensitization Effects by Bismuth Oxide Nanoparticles in Combination with Cisplatin for High Dose Rate Brachytherapy.

PURPOSE: The aim of this study was to investigate the potential of the synergetic triple therapeutic combination encompassing bismuth oxide nanoparticles (BiONPs), cisplatin (Cis), and high dose rate (HDR) brachytherapy with Ir-192 source in breast cancer and normal fibroblast cell line. METHODS: In vitro models of breast cancer cell lines (MCF-7, MDA-MB-231) and normal fibroblast cell line (NIH/3T3) were employed. Cellular localization and cytotoxicity studies were conducted prior to inspection on the radiosensitization effects and generation of reactive oxygen species (ROS) on three proposed radiosensitizers: BiONPs, Cis, and BiONPs-Cis combination (BC). The optimal, non-cytotoxic concentration of BiONPs (0.5 mM) and the 25% inhibitory concentration of Cis (1.30 µM) were applied. The radiosensitization effects were evaluated by using a 0.38 MeV Iridium-192 HDR brachytherapy source over a prescribed dose range of 0 Gy to 4 Gy. RESULTS: The cellular localization of BiONPs was visualized by light microscopy and accumulation of the BiONPs within the vicinity of the nuclear membrane was observed. Quantification of the sensitization enhancement ratio extrapolated from the survival curves indicates radiosensitization effects for MCF-7 and MDA-MB-231 when treated with BiONPs, Cis, and BC. However, NIH/3T3 cells exhibited contradictive behavior as it only reacted towards the BC combination. Nonetheless, the MCF-7 cell line loaded with BC shows the highest SER of 4.29. ROS production analysis, on the other hand, shows that Cis and BC radiosensitizers generated the highest free radicals in comparison to BiONPs alone. CONCLUSION: A BiONPs-Cis combination was unveiled as a novel approach that offers promising radiosensitization enhancement that will increase the efficiency of tumor control while preserving the normal tissue at a reduced dose. This data is the first precedent to prove the synergetic implication of BiONPs, Cis, and HDR brachytherapy that will be beneficial for future chemoradiotherapy strategies in cancer care.

Transfusion Medicine and Molecular Genetic Methods.

Transfusion procedures are always complicated by potential genetic mismatching between donor and recipient. Compatibility is determined by several major antigens, such as the ABO and Rhesus blood groups. Matching for other blood groups (Kell, Kidd, Duffy, and MNS), human platelet antigens, and human leukocyte antigens (HLAs) also contributes toward the successful transfusion outcomes, especially in multitransfused or highly immunized patients. All these antigens of tissue identity are highly polymorphic and thus present great challenges for finding suitable donors for transfusion patients. The ABO blood group and HLA markers are also the determinants of transplant compatibility, and mismatched antigens will cause graft rejection or graft-versus-host disease. Thus, a single and comprehensive registry covering all of the significant transfusion and transplantation antigens is expected to become an important tool in providing an efficient service capable of delivering safe blood and quickly locating matching organs/stem cells. This review article is intended as an accessible guide for physicians who care for transfusion-dependent patients. In particular, it serves to introduce the new molecular screening methods together with the biology of these systems, which underlies the tests.

Lymphocytic choriomeningitis virus infection of dendritic cells interferes with TLR-induced IL-12/IL-23 cytokine production in an IL-10 independent manner.

Dendritic cells produce IL-12 and IL-23 in response to viral and bacterial infection and these cytokines are responsible for successful pathogen clearance. How sequential viral and bacterial infections affect the production of IL-12 and IL-23 is currently not known. Our study demonstrates that in dendritic cells infected with Lymphocytic choriomeningitis virus (LCMV), TLR activation with bacterial PAMPs resulted in reduced IL-12 and IL-23 expression compared to non-infected cells. Furthermore, expression of other proinflammatory cytokines, TNF-α and IL-6, were not inhibited under these conditions. We discovered that TLR-induced phosphorylation of p38 was significantly inhibited in LCMV-infected cells. We detected enhanced expression of suppressor of cytokine signalling (SOCS)-3 and IL-10. Yet, neutralizing IL-10 did not restore IL-12/IL-23 expression. Taken together, these results show that virus infection interferes with the magnitude of TLR-mediated inflammatory responses by repressing specific cytokine expression.

Anti-viral Activities of Oroxylum indicum Extracts on Chikungunya Virus Infection.

Chikungunya virus (CHIKV) is a re-emerging mosquito-borne alphavirus that poses a threat to human worldwide. Driven by the lack of approved medication and vaccination, research on anti-Chikungunya agents has received great attention. In an effort to determine potential inhibitor of CHIKV, this study aimed at investigating the potential anti-viral activity of Oroxylum indicum extracts towards CHIKV-infected Vero cells. The virucidal, pre- and post-treatment effects of O. indicum were evaluated, using the maximum non-toxic dose of O. indicum methanol and aqueous extracts as determined by cytotoxicity assay. The viral inhibitory effect was assessed by the morphological changes of Vero cells and further confirmed by plaque assay. Both methanol and aqueous extracts of O. indicum had similar cytotoxicity in Vero cells. Interestingly, the virucidal effect of O. indicum aqueous extract revealed a significant reduction on the viral titre (p < 0.05). The prophylactic effect of aqueous extract was demonstrated when the pre-treated cells exhibited a significant anti-CHIKV activity (p < 0.05). However, methanol extract of this plant exerted an anti-viral activity against CHIKV only to a certain extent. Therefore, the aqueous extract of this plant has a potential to inhibit the virus and acts as prophylactic agent against CHIKV. Further studies however are needed to substantiate the finding and to determine the important compound of this plant as well as the mechanism of action in treating CHIKV infection.

IL-27-induced gene expression is downregulated in HIV-infected subjects.

OBJECTIVE: To characterize the effect of HIV infection on IL-27-induced gene expression. DESIGN: During HIV infection, cytokine expression and function become deregulated. IL-27 is an important modulator of inflammatory responses. Interestingly, IL-27 can inhibit HIV replication in T cells and monocytes, implicating IL-27 as a potential adjunct to anti-viral treatment. Our previous work demonstrated that circulating HIV may suppress IL-27 expression, therefore, this study, in continuation of our previous work, aimed to understand how HIV affects expression levels of the IL-27 receptor and downstream functions of IL-27. METHODS: Peripheral blood mononuclear cells (PBMC) were isolated from whole blood of HIV negative and HIV positive (viremic) individuals to assess IL-27-induced gene expression by flow cytometry and ELISA. PBMC were also processed for monocyte enrichment to assess IL-27 receptor expression by flow cytometry and real-time PCR. RESULTS: Expression of the IL-27 receptor subunit, gp130, was upregulated in response to IL-27 in HIV negative individuals, however, in HIV positive individuals, this IL-27 response was diminished. Furthermore, we observed downregulation of IL-27-induced IL-6, TNF-α, and IL-10 expression in HIV positive subjects. CONCLUSION: In HIV infection, IL-27-induced gene expression was impaired, indicating HIV-mediated dysregulation of IL-27 functions occurs during HIV infection. This study provides evidence for new viral pathogenic mechanisms contributing to the widespread impairment of immune responses observed in HIV pathogenesis.

Interleukin-23-induced interleukin-23 receptor subunit expression is mediated by the Janus kinase/signal transducer and activation of transcription pathway in human CD4 T cells.

Interleukin (IL)-23 plays a critical role in the development of the T helper (Th) cell response and is responsible for the maintenance of the IL-17 producing subset of Th cells, Th17. IL-23 is a heterodimeric cytokine composed of IL-23p19 and IL-12p40 subunits, and the signaling pathway for IL-23 involves 2 receptor chains: IL-12Rß1 and IL-23Rα. The IL-23 receptor complex is expressed on a number of cells, including natural killer cells, monocytes, macrophages, dendritic cells, and CD4 T cells. Currently, the molecular mechanisms governing expression of the IL-23 receptor chains, IL-23Rα and IL-12Rß1, are not well understood. Our results show that IL-23 induces upregulation of IL-23Rα and IL-12Rß1 expression in human CD4 T cells. Further, we demonstrate that inhibition of the Janus kinase/signal transducer and activation of transcription (JAK/STAT) pathway by SD-1029, a JAK2 inhibitor, 5'-deoxy-5'-(methylthio) adenosine, a STAT1 inhibitor, and STAT3 VII, a STAT3 inhibitor, were able to block IL-23-induced expression of IL-23 receptor subunits in the human SUPT-1 T cell line and in primary CD4 human T cells. Taken together, our results suggest a positive feedback regulation of the IL-23 receptor via IL-23-mediated activation of the JAK/STAT pathway.

Interleukin-27 induces a STAT1/3- and NF-kappaB-dependent proinflammatory cytokine profile in human monocytes.

IL-27 is a heterodimeric cytokine bridging innate and adaptive immunity by playing a role in the activation of naive T cells and in development of Th1 cells. Additionally, recent evidence supports a role for IL-27 in the activation of monocytic cells. Both pro-inflammatory and anti-inflammatory activities have been attributed to IL-27; however, the role played by IL-27 in the activation of human monocytic cells in terms of cytokine production has not been well described. Our results show that IL-27 is a strong inducer of proinflammatory cytokine and chemokine expression, including enhancement of IL-6, IP-10, MIP-1alpha, MIP-1beta, and TNF-alpha expression in human primary monocytes. Furthermore, we observed that IL-27-induced cytokine and chemokine production was mediated by STAT1, STAT3, and NF-kappaB activation. Understanding how IL-27 exerts its effects on monocytic cells will identify important molecular mechanisms in the regulation of immune responses, particularly in the modulation of monocyte activation.

Impact of HIV infection, highly active antiretroviral therapy, and hepatitis C coinfection on serum interleukin-27.

A newly described cytokine, interleukin-27 (IL-27), that activates naive CD4 T cells, has recently been shown to be an anti-HIV cytokine. However, the effect of HIV infection on IL-27 expression has not been characterized. We found that clinical characteristics, including HIV viral load, hepatitis C virus coinfection, and CD4 T cell counts, were associated with changes in serum IL-27. Overall, our results suggest circulating HIV may suppress IL-27, a critical concept in treatment development with this cytokine.

The IL-12 family of cytokines in infection, inflammation and autoimmune disorders.

Cytokines are critical coordinators of the immune response necessary for resolving bacterial and viral assaults on the immune system. In particular, the IL-12 family of cytokines are key players in the regulation of T cell responses. These responses are orchestrated by monocytes, macrophages, and dendritic cells which produce the members of the IL-12 family of cytokines in response to infection. IL-27 and IL-23 are two cytokines that are related to IL-12; these cytokines share homology at the subunit, receptor, and signalling levels. IL-12 is composed of p35 and p40 subunits, which, when combined together form the bioactive IL-12p70. IL-23 is composed of the IL-12p40 subunit as well as the IL-23p19 subunit, which shares homology with IL-12p35. IL-27 is composed of EBI3 and p28. These three cytokines activate similar members of the JAK/STAT signalling pathways as a result of homology in their receptor components. Production of these cytokines by activated monocytes, macrophages, and dendritic cells results in the activation and differentiation of T cells. In spite of their similarity, each of these cytokines has specific roles in the regulation of immune responses. IL-12 is required for the induction of IFN-gamma production, critical for the induction of Th1 cells. IL-27 has been shown to play a role in the induction of Th1 cells from naive T cells, whereas IL-23 has been demonstrated to play a key role in the induction of the newly described Th17 cells. Recently, a novel heterodimeric and anti-inflammatory cytokine composed of the IL-12p35 and EBI3 subunits has been identified as IL-35. The biological properties of the IL-12 family of cytokines, the signalling pathways mediated by these cytokines and their role in infection, inflammation, and autoimmune diseases will be the focus of this review.