Synthesis, Characterization, and Toxicity Assessment of Zinc Oxide-Doped Manganese Oxide Nanoparticles in a Macrophage Model.

The doping of engineered nanomaterials (ENMs) is a key tool for manipulating the properties of ENMs (e.g., electromagnetic, optical, etc.) for different therapeutic applications. However, adverse health outcomes and the cellular biointeraction of doped ENMs, compared to undoped counterparts, are not fully understood. Previously, we have shown that doping manganese oxide nanoparticles with ZnO (ZnO-MnO2 NPs) improved their catalytic properties. In this study, we assessed the toxicity of ZnO-MnO2 NPs in Raw 264.7 cells. NPs were prepared via an eco-friendly, co-precipitation method and characterized by several techniques, including transmission and scanning electron microscopy, X-ray diffraction, and Fourier transform infrared. The physicochemical properties of ZnO-MnO2 NPs, including size, morphology, and crystalline structure, were almost identical to MnO2 NPs. However, ZnO-MnO2 NPs showed slightly larger particle aggregates and negative charge in cell culture media. Exposure to ZnO-MnO2 NPs resulted in lower toxicity based on the cell viability and functional assay (phagocytosis) data. Exposure to both NPs resulted in the activation of the cell inflammatory response and the generation of reactive oxygen species (ROS). Despite this, exposure to ZnO-MnO2 NPs was associated with a lower toxicity profile, and it resulted in a higher ROS burst and the activation of the cell antioxidant system, hence indicating that MnO2 NP-induced toxicity is potentially mediated via other ROS-independent pathways. Furthermore, the cellular internalization of ZnO-MnO2 NPs was lower compared to MnO2 NPs, and this could explain the lower extent of toxicity of ZnO-MnO2 NPs and suggests Zn-driven ROS generation. Together, the findings of this report suggest that ZnO (1%) doping impacts cellular biointeraction and the consequent toxicological outcomes of MnO2 NPs in Raw 264.7 cells.

Whole-exome sequencing identifies cancer-associated variants of the endo-lysosomal ion transport channels in the Saudi population.

Background: Although national efforts are underway to document the genomic variability of the Saudi population relative to other populations, such variability remains largely unexplored. Genetic variability is known to impact the fate of cells and increase or decrease the risk of a variety of complex diseases including cancer forms. Therefore, the identification of variants associated with cancer susceptibility in Saudi population may protect individuals from cancer or aid in patient-tailored therapies. The endo-lysosomal ion transport genes responsible for cationic ion homeostasis within the cell. We screened 703 single-nucleotide polymorphisms (SNPs) of the endo-lysosomal ion transporter genes in the Saudi population and identified cancer-associated variants that have been reported in other populations. Methods: Utilizing previously derived local data of Whole-Exome Sequencing (WES), we examined SNPs of TPCN1, TPCN2, P2RX4, TRPM7, TRPV4, TRPV4, and TRPV6 genes. The SNPs were identified for those genes by our in-house database. We predicted the pathogenicity of these variants using in silico tools CADD, Polyphen-2, SIFT, PrimateAI, and FATHMM-XF. Then, we validated our findings by exploring the genetics database (VarSome, dbSNP NCB, OMIM, ClinVar, Ensembl, and GWAS Catalog) to further link cancer risk. Results: The WES database yielded 703 SNPs found in TPCN2, P2RX4, TRPM7, TRPV4, and TRPV6 genes in 1,144 subjects. The number of variants that were found to be common in our population was 150 SNPs. We identified 13 coding-region non-synonymous variants of the endo-lysosomal genes that were most common with a minor allele frequency (MAF) of ≥ 1 %. Twelve of these variants are rs2376558, rs3750965, rs61746574, rs35264875, rs3829241, rs72928978, rs25644, rs8042919, rs17881456, rs4987682, rs4987667, and rs4987657 that were classified as cancer-associated genes. Conclusion: Our study highlighted cancer-associated SNPs in the endo-lysosomal genes among Saudi individuals. The allelic frequencies on polymorphic variants confer susceptibility to complex diseases that are comparable to other populations. There is currently insufficient clinical data supporting the link between these SNPs and cancer risk in the Saudi population. Our data argues for initiating future cohort studies in which individuals with the identified SNPs are monitored and assessed for their likelihood of developing malignancies and therapy outcomes.

The Combination of Radiation with PARP Inhibition Enhances Senescence and Sensitivity to the Senolytic, Navitoclax, in Triple Negative Breast Tumor Cells.

Despite significant advances in the treatment of triple-negative breast cancer, this disease continues to pose a clinical challenge, with many patients ultimately suffering from relapse. Tumor cells that recover after entering into a state of senescence after chemotherapy or radiation have been shown to develop a more aggressive phenotype, and to contribute to disease recurrence. By combining the PARP inhibitor (PARPi), talazoparib, with radiation, senescence was enhanced in 4T1 and MDA-MB-231 triple-negative breast cancer cell lines (based on SA-ß-gal upregulation, increased expression of CDKN1A and the senescence-associated secretory phenotype (SASP) marker, IL6). Subsequent treatment of the radiation- and talazoparib-induced senescent 4T1 and MDA-MB231 cells with navitoclax (ABT-263) resulted in significant apoptotic cell death. In immunocompetent tumor-bearing mice, navitoclax exerted a modest growth inhibitory effect when used alone, but dramatically interfered with the recovery of 4T1-derived tumors induced into senescence with ionizing radiation and talazoparib. These findings support the potential utility of a senolytic strategy in combination with the radiotherapy/PARPi combination to mitigate the risk of disease recurrence in triple-negative breast cancer.

Protective role of Dodonaea viscosa extract against streptozotocin-induced hepatotoxicity and nephrotoxicity in rats.

Previous investigations have shown that D. viscosa herbal extract is often used to treat a variety of diseases. Therefore, the purpose of this study was to investigate any additional potential impacts on rat liver and kidney damage induced by diabetes. Streptozotocin (STZ) (60 mg/kg/day) was given as a single dosage to cause type 1 diabetes. After then, diabetic rats received oral doses of D. viscosa for four weeks at 150 and 300 mg/kg/day. Blood, liver, and kidney tissues were collected at the end of the treatment and examined. Analysis was made of the serum lipid profile, liver, and kidney functions, as well as blood biochemistry. Moreover, the levels of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), interleukin-1 beta (IL-1ß), prostaglandin E-2 (PGE-2), and nitric oxide (NO) were estimated in serum. In liver and kidney samples, thiobarbituric acid reactive substances (TBARs) and reduced glutathione (GSH), as well as the pro-inflammatory cytokines and enzymatic activities of glutathione peroxidase (GPx), glutathione reeducates (GR), glutathione-S-transferase (GST), catalase (CAT), and superoxide dismutase (SOD) were analyzed. Histological changes in liver and kidney cross-sections were also observed. Our findings demonstrated that D. viscosa dramatically decreased pro-inflammatory indicators in blood, kidney, and liver tissues as well as blood glucose, and restored insulin levels, and lipid profiles. Additionally, it significantly raises the antioxidant enzyme activity SOD, CAT, GPx, and GST, while significantly lowering TBARs levels. The above-mentioned biochemical changes that took place in tissues were further supported by histological alterations. These findings imply that D. viscosa protects against STZ-induced hyperglycemia, aberrant lipid synthesis, and oxidative stress and that these benefits may be mediated by interacting with various targets to increase the levels of antioxidant enzymes in the liver and kidneys. Its mode of action and safety for use as medicine against various metabolic problems caused by diabetes require more research.

Adverse Responses following Exposure to Subtoxic Concentrations of Zinc Oxide and Nickle Oxide Nanoparticles in the Raw 264.7 Cells.

The incorporation of engineered nanomaterials (ENMs) in biomedical and consumer products has been growing, leading to increased human exposure. Previous research was largely focused on studying direct ENM toxicity in unrealistic high-exposure settings. This could result in overlooking potential adverse responses at low and subtoxic exposure levels. This study investigated adverse cellular outcomes to subtoxic concentrations of zinc oxide (ZnONPs) or nickel oxide (NiONPs) nanoparticles in the Raw 264.7 cells, a macrophage-like cell model. Exposure to both nanoparticles resulted in a concentration-dependent reduction of cell viability. A subtoxic concentration of 6.25 µg/mL (i.e., no observed adverse effect level) was used in subsequent experiments. Exposure to both nanoparticles at subtoxic levels induced reactive oxygen species generation. Cellular internalization data demonstrated significant uptake of NiONPs, while there was minimal uptake of ZnONPs, suggesting a membrane-driven interaction. Although subtoxic exposure to both nanoparticles was not associated with cell activation (based on the expression of MHC-II and CD86 surface markers), it resulted in the modulation of the lipopolysaccharide-induced inflammatory response (TNFα and IL6), and cells exposed to ZnONPs had reduced cell phagocytic capacity. Furthermore, subtoxic exposure to the nanoparticles distinctly altered the levels of several cellular metabolites involved in cell bioenergetics. These findings suggest that exposure to ENMs at subtoxic levels may not be devoid of adverse health outcomes. This emphasizes the importance of establishing sensitive endpoints of exposure and toxicity beyond conventional toxicological testing.

GL67 lipid-based liposomal formulation for efficient siRNA delivery into human lung cancer cells.

The efficient delivery of small interfering RNA (siRNA) to the targeted cells significantly affects the regulation of the overexpressed proteins involved in the progression of several genetic diseases. SiRNA molecules in naked form suffer from low internalization across the cell membrane, high susceptibility to degradation by nuclease enzyme and low stability, which hinder their efficacy. Therefore, there is an urge to develop a delivery system that can protect siRNA from degradation and facilitate their uptake across the cell membrane. In this study, the cationic lipid (GL67) was exploited, in addition to DC-Chol and DOPE lipids, to design an efficient liposomal nanocarrier for siRNA delivery. The physiochemical characterizations demonstrated that the molar ratio of 3:1 has proper particle size measurements from 144 nm to 332 nm and zeta potential of -9 mV to 47 mV that depends on the ratio of the GL67 in the liposomal formulation. Gel retardation assay exhibited that increasing the percentage of GL67 in the formulations has a good impact on the encapsulation efficiency compared to DC-Chol. The optimal formulations of the 3:1 M ratio also showed high metabolic activity against A549 cells following a 24 h cell exposure. Flow cytometry findings showed that the highest GL67 lipid ratio (100 % GL67 and 0 % DC-Chol) had the highest percentage of cellular uptake. The lipoplex nanocarriers based on GL67 lipid could potentially influence treating genetic diseases owing to the high internalization efficiency and safety profile.

Effect of Autophagy Inhibitors on Radiosensitivity in DNA Repair-Proficient and -Deficient Glioma Cells.

Background and Objectives: The development of radioresistance is a fundamental barrier to successful glioblastoma therapy. Autophagy is thought to play a role in facilitating the DNA repair of DNA damage foci in radiation-exposed tumor cells, thus, potentially contributing to their restoration of proliferative capacity and development of resistance in vitro. However, the effect of autophagy inhibitors on DNA damage repair is not fully clear and requires further investigation. Materials and Methods: In this work, we utilized M059K (DNA-PKcs proficient) and M059J (DNA-PKcs deficient) glioma cell lines to investigate the role of autophagy inhibitors in the DNA repair of radiation-induced DNA damage. Cell viability following radiation was determined by trypan blue exclusion in both cell lines. Cell death and autophagy assays were performed to evaluate radiation-induced cell stress responses. DNA damage was measured as based on the intensity of phosphorylated γ-H2AX, a DNA double-stranded breaks (DSBs) marker, in the presence or absence of autophagy inhibitors. Results: The cell viability assay showed that M059J cells were more sensitive to the same dose of radiation (4 Gy) than M059K cells. This observation was accompanied by an elevation in γ-H2AX formation in M059J but not in M059K cells. In addition, the DAPI/TUNEL and Senescence-associated ß-galactosidase (SA-ß-gal) staining assays did not reveal significant differences in apoptosis and/or senescence induction in response to radiation, respectively, in either cell line. However, acridine orange staining demonstrated clear promotion of acidic vesicular organelles (AVOs) in both cell lines in response to 4 Gy radiation. Moreover, DNA damage marker levels were found to be elevated 72 h post-radiation when autophagy was inhibited by the lysosomotropic agent bafilomycin A1 (BafA1) or the PI3K inhibitor 3-methyl adenine (3-MA) in M059K cells. Conclusions: The extent of the DNA damage response remained high in the DNA-PKcs deficient cells following exposure to radiation, indicating their inability to repair the newly formed DNA-DSBs. On the other hand, radioresistant M059K cells showed more DNA damage response only when autophagy inhibitors were used with radiation, suggesting that the combination of autophagy inhibitors with radiation may interfere with DNA repair efficiency.

CC chemokine receptor 5 antagonist alleviates inflammation by regulating IFN-γ/IL-10 and STAT4/Smad3 signaling in a mouse model of autoimmune encephalomyelitis.

Multiple sclerosis (MS) is an immunopathological disease that causes demyelination and recurrent episodes of T cell-mediated immune attack in the central nervous system. Experimental autoimmune encephalomyelitis (EAE) is a well-established mouse model of MS. The roles of T cells in MS/EAE have been well investigated, but little is known about the role of CCR5+ cells. In the present study, we investigated whether treatment with DAPTA, a selective CCR5 antagonist, could modulate the progression of EAE in the SJL/J mice. EAE mice were treated with DAPTA (0.01 mg/kg) intraperitoneally daily from day 14 to day 42, and the clinical scores were evaluated. We further investigated the effects of DAPTA on IFN-γ-, TGF-ß-, IL-10-, IL-17A-, IL-22-, T-bet, STAT4-, RORγT-, AhR-, Smad3-, and Foxp3-expressing CCR5+ spleen cells using flow cytometry analysis. We further explored the effects of DAPTA on mRNA/protein expression of IFN-γ, IL-10, IL-17A, IL-22, TGF-ß, T-bet, STAT4, RORγT, AhR, Foxp3, and NF-H in the brain tissue. The severity of clinical scores decreased in DAPTA-treated EAE mice as compared to that in the EAE control mice. Moreover, the percentage of CCR5+IFN-γ+, CCR5+T-bet+, CCR5+STAT4+, CCR5+IL-17A+, CCR5+RORγt+, CCR5+IL-22+, and CCR5+AhR+ cells decreased while CCR5+TGF-ß+, CCR5+IL-10+, CCR5+Smad3+, and CCR5+Foxp3+ increased in DAPTA-treated EAE mice. Furthermore, DAPTA treatment significantly mitigated the EAE-induced expression of T-bet, STAT4, IL-17A, RORγT, IL-22, and AhR but upregulated Foxp3, IL-10, and NF-H expression in the brain tissue. Taken together, our data demonstrated that DAPTA could ameliorate EAE progression through the downregulation of the inflammation-related cytokines and transcription factors signaling, which may be useful for the clinical therapy of MS.

Metformin attenuates V-domain Ig suppressor of T-cell activation through the aryl hydrocarbon receptor pathway in Melanoma: In Vivo and In Vitro Studies.

Melanoma is an aggressive skin cancer with a high rate of metastasis to other organs. Recent studies specified the overexpression of V-domain Ig suppressor of T-cell activation (VISTA) and Aryl Hydrocarbon Receptor (AHR) in melanoma. Metformin shows anti-tumor activities in several cancer types. However, the mechanism is unclear. This study aims to investigate the inhibitory effect of metformin on VISTA via AHR in melanoma cells (CHL-1, B16) and animal models. VISTA and AHR levels were assessed by qPCR, Western blot, immunofluorescence microscope, flow cytometry, and immunohistochemistry. Here, metformin significantly decreased VISTA and AHR levels in vitro and in vivo. Furthermore, metformin inhibited all AHR-regulated genes. VISTA levels were dramatically inhibited by AHR modulations using shRNA and αNF, confirming the central role of AHR in VISTA. Finally, melanoma cells were xenografted in C57BL/6 and nude mice. Metformin significantly reduced the tumor volume and growth rate. Likewise, VISTA and AHR-regulated protein levels were suppressed in both models. These findings demonstrate for the first time that VISTA is suppressed by metformin and identified a new regulatory mechanism through AHR. The data suggest that metformin could be a new potential therapeutic strategy to treat melanoma patients combined with targeted immune checkpoint inhibitors.

Aptamer­based therapy for targeting key mediators of cancer metastasis (Review).

Cancer­related deaths remain a challenging and devastating obstacle to defeat despite the tremendous advances in cancer treatment. Cancer metastasis is the major cause of these cancer­related deaths. Metastasis involves sequential steps during cancer cells' journey to a new site. These steps are coordinately regulated by specific intracellular regulators and cellular interactions between the cancer cells and the supporting microenvironment of the different organs. The development of aptamer­based therapeutics is a promising strategy to fight cancer metastasis as it holds potential advantages. Oligonucleotide and peptide aptamers are short sequences of single­stranded nucleic acids or amino acids, respectively, that target proteins, genetic materials, and cells. Antimetastatic aptamer­based therapeutics exert their pharmacological effect by direct interaction with the signaling pathways inside the cancer cells or the communications between cancer cells and the tumor microenvironment. In addition, aptamers have been utilized as a guiding ligand to deliver a therapeutic moiety to cancer cells or the supporting microenvironment. The selected aptamer possesses high specificity since it is designed to recognize and interact with its target. This review summarizes recent advances in the development of aptamer­based therapeutics targeting mediators of cancer metastasis. In addition, potential opportunities are discussed to inspire researchers in the field to develop novel aptamer­based antimetastatic treatments.

Sorafenib, rapamycin, and venetoclax attenuate doxorubicin-induced senescence and promote apoptosis in HCT116 cells.

Emerging evidence has shown that the therapy-induced senescent growth arrest in cancer cells is of durable nature whereby a subset of cells can reinstate proliferative capacity. Promising new drugs named senolytics selectively target senescent cells and commit them into apoptosis. Accordingly, senolytics have been proposed as adjuvant cancer treatment to cull senescent tumor cells, and thus, screening for agents that exhibit senolytic properties is highly warranted. Our study aimed to investigate three agents, sorafenib, rapamycin, and venetoclax for their senolytic potential in doxorubicin-induced senescence in HCT116 cells. HCT116 cells were treated with one of the three agents, sorafenib (5 µM), rapamycin (100 nM), or venetoclax (10 µM), in the absence or presence of doxorubicin (1 µM). Senescence was evaluated using microscopy-based and flow cytometry-based Senescence-associated-ß-galactosidase staining (SA-ß-gal), while apoptosis was assessed using annexin V-FITC/PI, and Muse caspase-3/-7 activity assays. We screened for potential genes through which the three drugs exerted senolytic-like action using the Human Cancer Pathway Finder PCR array. The three agents reduced doxorubicin-induced senescent cell subpopulations and significantly enhanced the apoptotic effect of doxorubicin compared with those treated only with doxorubicin. The senescence genes IGFBP5 and BMI1 and the apoptosis genes CASP7 and CASP9 emerged as candidate genes through which the three drugs exhibited senolytic-like properties. These results suggest that the attenuation of doxorubicin-induced senescence might have shifted HCT116 cells to apoptosis by exposure to the tested pharmacological agents. Our work argues for the use of senolytics to reduce senescence-mediated resistance in tumor cells and to enhance chemotherapy efficacy.

A novel cyclic dinuclear gold(I) complex induces anticancer activity via an oxidative stress-mediated intrinsic apoptotic pathway in MDA-MB-231 cancer cells.

A new dinuclear cyclic gold(I) complex [Au2(DCyPA)2](PF6)2, 1, based on bis[2-(dicyclohexylphosphano)ethyl]amine (DCyPA) has been synthesized and characterized by elemental analysis, IR and NMR spectroscopy, and X-ray crystallography. In the dinuclear complex cation [Au2(DCyPA)2]2+, the two gold(I) ions are bridged by the ligand bis[2-(dicyclohexylphosphano)ethyl]amine (DCyPA) giving rise to a 16-membered ring centrosymmetric metallacycle. The cytotoxicity of the complex was evaluated against the triple-negative human breast cancer cells MDA-MB-231. In order to understand the mechanism of the cytotoxic behavior, a variety of assays, including Annexin V-FITC/Propidium iodide double staining, ROS production, and mitochondrial membrane potential and migration assays were carried out. The results indicated that complex 1 induced cytotoxicity via an oxidative stress-mediated intrinsic apoptotic pathway in MDA-MB-231 cancer cells.

In vivo and in vitro studies evaluating the chemopreventive effect of metformin on the aryl hydrocarbon receptor-mediated breast carcinogenesis.

Metformin (MET) is a clinically used anti-hyperglycemic agent that shows activities against chemically-induced animal models of cancer. A study from our laboratory showed that MET protectes against 7, 12-dimethylbenz[a]anthracene (DMBA)-induced carcinogenesis in vitro human non-cancerous epithelial breast cells (MCF10A) via activation of the aryl hydrocarbon receptor (AhR). However, it is unclear whether MET can prevent the initiation of breast carcinogenesis in an in vivo rat model of AhR-induced breast carcinogenesis. Therefore, the main aims of this study are to examine the effect of MET on protecting against rat breast carcinogenesis induced by DMBA and to explore whether this effect is medicated through the AhR pathway. In this study, treatment of female rats with DMBA initiated breast carcinogenesis though inhibiting apoptosis and tumor suppressor genes while inducing oxidative DNA damage and cell cycle proliferative markers. This effect was associated with activation of AhR and its downstream target genes; cytochrome P4501A1 (CYP1A1) and CYP1B1. Importantly, MET treatment protected against DMBA-induced breast carcinogenesis by restoring DMBA effects on apoptosis, tumor suppressor genes, DNA damage, and cell proliferation. Mechanistically using in vitro human breast cancer MCF-7 cells, MET inhibited breast cancer stem cells spheroids formation and development by DMBA, which was accompanied by a proportional inhibition in CYP1A1 gene expression. In conclusion, the study reports evidence that MET is an effective chemopreventive therapy for breast cancer by inhibiting the activation of CYP1A1/CYP1B1 pathway in vivo rat model.

Protective effect of valsartan against doxorubicin-induced cardiotoxicity: Histopathology and metabolomics in vivo study.

Doxorubicin (DOX) treatment has been associated with cardiotoxicity. Therefore, it is crucial to search for a therapeutic that can effectively mitigate DOX-induced cardiotoxicity. This study was conducted to investigate the protective effects of valsartan (VAL) against DOX-induced cardiotoxicity. Sprague-Dawley rats were divided into four treatment groups: Group I: Control, Group II: VAL (30 mg/kg, ip), Group III: DOX (15 mg/kg, ip), and Group IV: VAL + DOX (30 + 15 mg/kg, ip). All groups were treated every other day for 14 days. Blood was isolated for biochemical and metabolomics studies, and sections of the heart were also analyzed for histopathological and immunohistochemical alterations to detect changes in P53, BAX, BCL-2, and P62 expression. The combination of VAL + DOX resulted in a marked decrease in cardiac biomarker enzymes (aminotransferase and creatine phosphokinase) compared to DOX monotherapy. In addition, the histopathological examination of the VAL + DOX combination revealed a low percentage of fibrosis and inflammation. Immunohistochemical expression of p53 and BAX was significantly reduced, whereas BCL-2 expression was significantly increased in the VAL + DOX treatment group compared to DOX monotherapy. Also, the combination of VAL + DOX reverses the negative effect of DOX on nuclear p62 expression. Analysis of serum metabolites showed that DOX monotherapy reduced the number of several amino acids, whereas the combination of VAL + DOX restored these metabolic pathways. This study revealed the potential cardioprotective effect of VAL, which may provide novel and promising approaches for managing cardiotoxicity induced by DOX.

Inhibition of interleukin-2-inducible T-cell kinase causes reduction in imiquimod-induced psoriasiform inflammation through reduction of Th17 cells and enhancement of Treg cells in mice.

Psoriasis is a debilitating chronic skin disease with a worldwide prevalence. Its main features include well-marked silvery scales on the skin of hands and feet and back which arise due to hyperproliferation of keratinocytes and infiltration of immune cells in the skin. Multiple interactions exist between adaptive immune cells such as T cells and innate immune cells such as neutrophils and macrophages which are key players in the pathogenesis of psoriasis. Interleukin-2-inducible T-cell kinase (ITK) plays a key role in Th17 cell development through control of several transcription factors. ITK has been shown to control NFATc1, NFkB and STAT3 in CD4+ T cells. Effect of ITK inhibitor in imiquimod (IMQ)-induced psoriasiform inflammation remains to be explored. In the current examination, role of ITK signaling and its inhibition blockade were evaluated on NFATc1, NFkB and STAT3, IL-17A, TNF-α, IFN-γ, Foxp3, IL-10 in CD4+ T cells in IMQ model. Our data display that ITK signaling is involved in IMQ-induced psoriatic inflammation as paralleled by enhancement of p-ITK, NFATc1, p-NFkB and p-STAT3 in CD4+ T cells. It was associated with enhancement of Th17/Th1 cells and neutrophilic inflammation in the skin. Preventive treatment with ITK inhibitor led to a reduction in Th17/Th1 cells and enhancement of Treg cells. Overall, this study suggests that ITK signaling is an important modulator of transcription factor signaling in CD4+ T cells which is associated with Th17/Th1 cells and psoriasiform inflammation in mice. ITK signaling blockade could be a therapeutic target for the treatment of psoriatic inflammation.

3-Aminobenzamide alleviates elevated DNA damage and DNA methylation in a BTBR T+Itpr3tf/J mouse model of autism by enhancing repair gene expression.

Little is known about genetic and epigenetic alterations in autism spectrum disorder. Moreover, the efficiency of DNA repair in autism must be improved to correct these alterations. We examined whether 3-aminobenzamide (3-AB) could reverse these alterations. We conducted experiments to clarify the molecular mechanism underlying these ameliorations. An assessment of genetic and epigenetic alterations by a modified comet assay showed elevated levels of oxidative DNA strand breaks and DNA hypermethylation in BTBR T+Itpr3tf/J (BTBR) mice used as a model of autism. Oxidative DNA strand breaks and DNA methylation were further quantified fluorometrically, and the results showed similar changes. Conversely, 3-AB treated BTBR mice showed a significant reduction in these alterations compared with untreated mice. The expressions of 43 genes involved in DNA repair were altered in BTBR mice. RT2 Profiler PCR Array revealed significantly altered expression of seven genes, which was confirmed by RT-PCR analyses. 3-AB treatment relieved these disturbances and significantly improved Ogg1 and Rad1 up-regulation. Moreover, autism-like behaviors were also mitigated in BTBR animals by 3-AB treatment without alterations in locomotor activities. The simultaneous effects of reduced DNA damage and DNA methylation levels as well as the regulation of repair gene expression indicate the potential of 3-AB as a therapeutic agent to decrease the levels of DNA damage and DNA methylation in autistic patients. The current data may help in the development of therapies that ultimately provide a better quality of life for individuals suffering from autism.

Ubiquitous plasticizer, Di-(2-ethylhexyl) phthalate enhances existing inflammatory profile in monocytes of children with autism.

Genetic as well as environmental factors are believed to play a significant role in the pathogenesis and progression of autism spectrum disorder (ASD). Phthalates are ubiquitous environmental contaminants as they are used plasticizers in several household/industrial products such as vinyl flooring, plastic toys, and cosmetic products. One of the plasticizers that is quite prevalent in these products is di-2-ethylhexyl phthalate (DEHP) which can cause human exposure via dermal/inhalation/ingestion routes. DEHP and its metabolites are associated with behavioral dysregulations and reported to be increased in systemic circulation of ASD children. DEHP is reported to cause upregulation of several inflammatory cytokines in different cells/tissues, however its role in inflammatory signaling of ASD monocytes has not been investigated earlier. Therefore, this study evaluated the effects of DEHP (at 5 µM final concentration for 24 h) on inflammatory profile (NFkB, STAT3, IL-6, TNF-α, IL-1ß) in monocytes of ASD subjects and typically developing control (TDC) children. Our data show that DEHP upregulates NFkB/STAT3 expression which is associated with increased inflammatory profile in monocytes of ASD and TDC subjects, however its effect is much greater in magnitude in the former group. This was confirmed by utilization of NFkB inhibitor, PDTC and STAT3 inhibitor, Stattic which caused reduction in inflammatory cytokines from DEHP-treated monocytes in ASD group. In short, DEHP causes further elevation in inflammatory signaling in ASD monocytes which could be due to existing inflammation in this group. These data suggest that use of plasticizers such as DEHP should be minimized in order to avoid their potential effects on immune dysfunction associated with ASD.

Elevated expression of toll-like receptor 4 is associated with NADPH oxidase-induced oxidative stress in B cells of children with autism.

Autism spectrum disorder (ASD) is a childhood disorder with neurodevelopmental dysfunction which manifests as impairment in social behavior and communication skills. B cells play an important role in immune dysfunction where toll-like receptor 4 (TLR4) may contribute through oxidative inflammatory process. TLR4 related signaling and oxidative stress have been reported in the periphery of ASD subjects, however it has not been evaluated in peripheral B cells of ASD subjects and compared with typically developing control (TDC) children. This study evaluated TLR4 expression and related signaling [Bruton's tyrosine kinase (BTK), spleen tyrosine kinase (SYK), NF-kB, NADPH oxidase (NOX2), nitrotyrosine, superoxide dismutase (SOD)] in ASD and TDC subjects. Current investigation in B cells shows that ASD subjects have increased TLR4 expression and oxidative stress as exhibited by upregulated NOX2 and nitrotyrosine expression as compared to TDC subjects. B cell relevant pathways, BTK/SYK/NF-kB were also upregulated in B cells of ASD group. Treatment with TLR4 agonist, LPS led to upregulation of NOX2 and nitrotyrosine in B cells of ASD whereas it had no significant effect on TDC subjects. Treatment with NF-kB inhibitor caused inhibition of LPS-induced upregulation of NOX2 and nitrotyrosine in B cells of ASD. Therefore, current investigation proposes the notion that TLR4 expression is elevated in B cells which is associated with increased NF-kB signaling and oxidant stress in ASD subjects. In short, peripheral B cells could contribute to systemic oxidative inflammation and contribute to the immune dysfunction in ASD.

Liraglutide attenuates gefitinib-induced cardiotoxicity and promotes cardioprotection through the regulation of MAPK/NF-κB signaling pathways.

Gefitinib is an effective treatment for patients with locally advanced non-small cell lung cancer. However, it is associated with cardiotoxicity that can limit its clinical use. Liraglutide, a glucagon-like peptide 1 receptor agonist, showed potent cardioprotective effects with the mechanism is yet to be elucidated. Therefore, this study aimed to determine the efficiency of liraglutide in protecting the heart from damage induced by gefitinib. Adult male Wistar rats were randomly divided into control group, liraglutide group (200 µg/kg by i.p. injection), gefitinib group (30 mg/kg orally) and liraglutide plus gefitinib group. After 28 days, blood and tissue samples were collected for histopathological, biochemical, gene and protein analysis. We demonstrated that gefitinib treatment (30 mg/kg) resulted in cardiac damage as evidenced by histopathological studies. Furthermore, serum Creatine kinase-MB (CK-MB), N-terminal pro B-type natriuretic peptide (NT-proBNP) and cardiac Troponin-I (cTnI) were markedly elevated in gefitinib group. Pretreatment with liraglutide (200 µg/kg), however, restored the elevation in serum markers and diminished gefitinib-induced cardiac damage. Moreover, liraglutide improved the gene and protein levels of anti-oxidant (superoxide dismutase) and decreased the oxidative stress marker (NF-κB). Mechanistically, liraglutide offered protection through upregulation of the survival kinases (ERK1/2 and Akt) and downregulation of stress-activated kinases (JNK and P38). In this study, we provide evidence that liraglutide protects the heart from gefitinib-induced cardiac damage through its anti-oxidant property and through the activation of survival kinases.

The histamine-4 receptor antagonist JNJ7777120 prevents immune abnormalities by inhibiting RORγt/T-bet transcription factor signaling pathways in BTBR T+ Itpr3tf/J mice exposed to gamma rays.

Autism is a neurodevelopmental disorder characterized by deficits and qualitative impairments in communication and implicit skill learning. Its prevalence is higher than previous estimates, and treatments have limited efficacy and are costly. Here, we assessed the therapeutic potential of JNJ77777120 (JNJ), a histamine-4 receptor (H4R) antagonist, using BTBR T+ Itpr3tf/J (BTBR) mice, a confirmed model of autism, and C57BL/6J (C57) mice, a commonly chosen reference strain. We first examined the effects of JNJ treatment on BTBR mice exposed to gamma-rays (irradiation-exposed) using a three-chambered apparatus. We further investigated the possible molecular mechanisms through which JNJ administration modulates IL-17A-, RORγT-, IL-22-, T-bet-, STAT3-, ICOS-, and Foxp3-producing CD8+ T cells in the spleens of irradiation-exposed BTBR mice. The effects of JNJ administration on the mRNA and protein expression of IL-17A, RORγT, IL-22, T-bet, STAT-3, pSTAT3, IL-10, and Foxp3 in brain tissue were also explored. Results showed that JNJ treatment with irradiation exposure increased social interactions in BTBR mice compared to that in irradiation-exposed BTBR mice. Additionally, JNJ-treated and irradiation-exposed BTBR mice exhibited decreases in IL-17A-, RORγT-, IL-22-, T-bet-, and STAT3-producing CD8+ T cells and increases in ICOS- and Foxp3-producing CD8+ T cells. Moreover, JNJ treatment and irradiation exposure in BTBR mice regulated the mRNA and protein expression levels of IL-17A, RORγT, IL-22, T-bet, STAT3, pSTAT-3, IL-10, and Foxp3 in the brain tissue. These results suggest that JNJ is useful for the treatment of autism, as this H4R antagonist could block inflammatory cytokine production and transcription factor signaling.