Ribonucleic Acid Levels Research Articles

M1 macrophage-derived exosome-encapsulated cisplatin can enhance its anti-lung cancer effect.

The aim of this study was to study the efficacy of cisplatin-loaded M1 macrophage secreted-exosomes (DDP-M1-Exos) in enhancing DDP antitumor effect. M1-Exos were first extracted using density gradient centrifugation, and the DDP-M1-Exos system was established via electroporation. Then, the morphology, particle size and maker proteins of the DDP-M1-Exos system were assessed using transmission electron microscope, DLS and Western blotting (WB), respectively. The uptake of Exos by mouse Lewis lung cancer (LLC) cells was observed under a confocal laser scanning microscope, and the influence of the DDP-M1-Exos system on the viability of Lewis cells was determined using methyl thiazolyl tetrazolium assay and 4',6-diamidino-2-phenylindole staining. Besides, its impacts on the messenger ribonucleic acid (mRNA) levels and protein expression levels of B-cell lymphoma 2 (Bcl-2), Bcl-2 associated X protein (Bax) and Caspase-3 in LLC cells were examined using quantitative real-time PCR (qRT-PCR) and WB, respectively. Finally, with the LLC-bearing mouse model as the object of study, the efficacy was evaluated using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) and hematoxylin-eosin staining staining in vivo. Saucer-shaped Exos with a bilayer membrane, uniform particle size and highly expressed CD81, Alix and HSP70 were successfully isolated from M1 macrophages via density gradient centrifugation, and they were absorbed by LLC cells. Meanwhile, the DDP-M1-Exos drug delivery system was successfully prepared using electroporation technique. Compared with Control group, DDP and DDP-M1-Exos groups exhibited a decrease in the proliferation rate of mouse LLC cells and an increase in their apoptosis rate (*P<0.05), and the apoptosis rate in DDP-M1-Exos group was substantially higher than that in DDP group (#P<0.01). According to the qRT-PCR and WB results, DDP-M1-Exos up-regulated Bax and Caspase-3 in the apoptosis signaling pathway to induce tumor cell apoptosis, thereby resisting tumors. Moreover, through in-vivo experiments, it was found that M1-Exos alone had a potential to suppress tumor growth, and that the carrier of M1-Exos could not only kill tumor cells, but also encapsulate DDP to enhance its anti-lung cancer effect. The DDP-M1-Exos drug delivery system is prepared using Exos extracted via density gradient centrifugation by electroporation, and the drug-loaded Exos are able to effectively inhibit the proliferation of tumor cells and induce their apoptosis, exerting an antitumor effect.

Human Sperm Cells After Purification Using SCLB Can Be Stored at 4o, -20o, or -80oC Before Small RNA Isolation

There have been many studies about pre-analysis for sperm RNA examination which compared sperm purificationmethods, RNA isolation methods, sequencing methods, and semen storage before analysis. However, there is a lack ofstudies that determine the ideal storage temperature after sperm cell purification before RNA analysis, especially small RNAanalysis. The aim of this study was to determine the preferred storage temperature for human sperm cells after spermpurification using Somatic Cell Lysis Buffer (SCLB) before sperm small ribonucleic acid (RNA) isolation and analysis. Thisstudy was a true laboratory experiment using the post-test only control group design. The samples were 13 fresh humansemen that has been purified using SCLB. The sperm cells were then diluted and divided into four aliquots with differenttreatments. The first aliquot that served as a control group was immediately purified while the other three aliquots were0 0 0 stored for seven days at different temperatures as follows: 4 C, -20 , and -80 C. After the small RNA isolation, RNA levelbetween each group was compared. Micro volume spectrophotometer measured RNA level. The median of small RNA6 yields of the control group was 49.8 (5.33-522.46) ng/10 sperm cells. There was no significant difference in median of smallRNA yields of the control group and that of other groups. The median of the other groups with storage temperature0 0 0 6 of 4 C, -20 , and -80 C was 41.09 (7.03-1448.31), 65.95 (7.99-301.16), and 76.42 (10.45-434.25) ng/10 sperm cells,respectively (p-value= 0.314; α=5%). This condition suggested that after purification using SCLB, human sperm cells can be0 0 0 stored at temperatures of 4 C, -20 , or -80 C for seven days, depending on each laboratory facility.

Effect of Emodin on Coxsackievirus B3m-Mediated Encephalitis in Hand, Foot, and Mouth Disease by Inhibiting Toll-Like Receptor 3 Pathway In Vitro and In Vivo.

Encephalitis in hand, foot, and mouth disease (HFMD) is a serious threat to children's health and life. Toll-like receptor 3 (TLR3) is an innate immune-recognition receptor that can recognize virus and initiate innate immune responses. Emodin has the effects of anti-inflammatory and regulating immune function, but the mechanism is not very clear. Cells and mice were pretreated with coxsackievirus B3m (CVB3) and treated with emodin. The messenger ribonucleic acid (mRNA) and protein levels of TLR3 and downstream molecules were detected by quantitative real-time polymearse chain reaction and western blotting analysis, respectively. TLR3 expression was also downregulated by anti-TLR3 antibody (TLR3Ab) or small interfering RNA (siRNA). Pathological changes were assessed with hematoxylin and eosin staining. Immunohistochemistry was used to examine the expression of TLR3 in brain tissues. The expression of interleukin (IL)-6, nuclear factor (NF)-κB, and interferon (IFN)-β in serum were tested with enzyme-linked immunosorbent assay. Emodin decreased the mRNA and protein levels of TLR3 and downstream molecules in vitro and in vivo. After downregulating TLR3 using anti-TLR3Ab or siRNA, emodin could still decrease the mRNA and protein levels of TLR3 and downstream molecules. Emodin also displayed notable effects on pathology, TLR3 protein in brain tissues, and expression of IL-6, NF-κB, IFN-β, in serum. Emodin exerts a protective effect in CVB3-mediated encephalitis in HFMD by inhibiting the TLR3 pathway.

Ten Representative Saponins on Tissue Factor Expression in Human Monocytes: Structure–Activity Relationships and Molecular Docking

Saponins have significant bioactivities in treating cardiovascular disease. Whereas there is a lack of in-depth knowledge about how saponins prevent cardiovascular disease. Tissue factor (TF) is the major initiator of the coagulation cascade and plays an important role in hemostasis and thrombosis. However structure–activity relationships (SARs) of saponins inhibiting TF activity have not been discussed in detail at present. To further clarify the relationships between saponins and TF, in this study, 10 representative saponins were selected to study the inhibitory effect on TF procoagulant activity of monocytes by an improved chromogenic substrate method, and the possible SARs were preliminarily analyzed. Furthermore, molecular docking analysis suggested that 4 representative saponins had a good affinity with TF/FVIIa. In addition, a representative saponin, ruscogenin, decreased both messenger ribonucleic acid and protein levels of TF in human monocytes partly due to its downregulation of nuclear factor kappa-light-chain-enhancer of activated B cells and c-Jun N-terminal kinase pathways. In conclusion, this study provides further explanation for the cardiovascular protection of saponins, and the analysis of SARs between inhibiting TF activity and saponins will be helpful to explore the therapeutic TF inhibitors.

Human Case Infected With Babesia venatorum: A 5-Year Follow-Up Study

BackgroundHuman babesiosis is a common zoonosis caused by Babesia and is attracting an increasing concern worldwide. The natural course of babesiosis infection and how the human immune system changes during the course of babesiosis infection are not clear.MethodsWe followed up 1 case infected with Babesia venatorum for 5 years. The patient was immune-intact and received no standard treatment. Clinical data were obtained from medical records. Microbiological tests, ribonucleic acid (RNA) sequence, and serum cytokines and chemokines were detected at different time points.ResultsThe patient was confirmed as B venatorum infection based on his tick-bite history, clinical manifestations, and positive results of microbiological tests. The parasitemia of the patient persisted for approximately 2 months. With flu-like symptoms aggravating, most cytokines and chemokines in RNA and protein levels increased progressively and reached the peak when fever occurred; and their concentrations decreased to baseline during the same time as clearance of babesia parasites.Conclusions Babesia venatorum infection could take a mild self-limited course in immune-intact individuals. The natural changes of most cytokines and chemokines demonstrated very similar trends, which correlated with blood parasitemia and clinical manifestations. Cytokine profiles involving multiple inflammatory cytokines might be a good indicator of babesia infection.

Estradiol Potentiates But Is Not Essential for Prolactin-Induced Suppression of Luteinizing Hormone Pulses in Female Rats.

Hyperprolactinemia causes infertility by suppressing gonadotropin-releasing hormone (GnRH) and luteinizing hormone (LH) secretion. Because effects of prolactin (PRL) on the hypothalamus usually require estradiol (E2), we investigated the role of E2 in PRL-induced suppression of LH pulses. Ovariectomized (OVX) rats treated with oil or E2 (OVX + E2) received a subcutaneous injection of ovine PRL (oPRL) 30 minutes before serial measurement of LH in the tail blood by enzyme-linked immunosorbent assay. E2 reduced pulsatile LH secretion. oPRL at 1.5 mg/kg further reduced LH pulse frequency in OVX + E2 but had no effect in OVX rats. The higher dose of 6-mg/kg oPRL decreased LH pulse frequency in both OVX and OVX + E2 rats, whereas pulse amplitude and mean LH levels were lowered only in OVX + E2 rats. Kisspeptin immunoreactivity and Kiss1 messenger ribonucleic acid (mRNA) levels were decreased in the arcuate nucleus (ARC) of OVX + E2 rats. oPRL decreased both kisspeptin peptide and gene expression in the ARC of OVX rats but did not alter the already low levels in OVX + E2 rats. In the anteroventral periventricular nucleus, oPRL did not change kisspeptin immunoreactivity and, paradoxically, increased Kiss1 mRNA only in OVX + E2 rats. Moreover, oPRL effectively reduced Gnrh expression regardless of E2 treatment. In this study we used tail-tip blood sampling to determine the acute effect of PRL on LH pulsatility in female rats. Our findings characterize the role of E2 in the PRL modulation of hypothalamic components of the gonadal axis and LH release, demonstrating that E2 potentiates but is not essential for the suppression of pulsatile LH secretion caused by hyperprolactinemia.

Effect of TGF-β1 on myocardial cell apoptosis in rats with acute myocardial infarction via MAPK signaling pathway.

Transforming growth factor beta 1 (TGF-β1) can promote myocyte hypertrophy, thus playing an important role in ventricular remodeling after myocardial infarction (MI). In this study, the model of MI was established in rats through ligating the left anterior descending coronary artery. Subsequently, the messenger ribonucleic acid (mRNA) and protein expression levels of TGF-β1 in myocardial cells in both model group and sham operation group were determined. The effects of TGF-β1 treatment on myocardial cell apoptosis in MI rats were explored. Moreover, the changes of mitogen-activated protein kinase (MAPK) signaling pathway in rats with acute MI were verified. In addition, the protein expressions of phosphorylated-MAPK kinases 3/6 (p-MKK3/6) and MKK3/6 in myocardial cells of the two groups were analyzed. The mRNA and protein expression levels of TGF-β1 in myocardial cells of acute MI rats were significantly higher than those in the sham operation group (p<0.01). After treatment with TGF-β1, the expression level of B-cell lymphoma 2 (Bcl-2) associated X protein (Bax) was obviously down-regulated. The Bax/Bcl-2 ratio was notably lower than that in control group (p<0.01). Meanwhile, the proportion of apoptotic cells decreased remarkably (p<0.01). In the model group, no evident change was observed in the protein expression level of MKK3/6, whereas the levels of p-MKK3/6 were prominently up-regulated (p<0.01). TGF-β1 can activate MKK3/6 in the MAPK signaling pathway to resist the apoptosis of myocardial cells in acute MI rats.

Effect of strontium ranelate on rabbits with steroid-induced osteonecrosis of femoral head through TGF-β1/BMP2 pathway.

To study the effect of strontium ranelate (SR) on steroid-induced osteonecrosis of the femoral head (SIONFH) in rabbits and its regulatory mechanism. The ONFH model was established in 30 rabbits using steroid and they were randomly divided into Control group, Model group, and SR group. After SR intervention, the rabbits were sacrificed and sampled. The pathological injury of the femoral head in each group was detected via hematoxylin-eosin (HE) staining, the level of vascular endothelial growth factor (VEGF) in the femoral head in each group was detected via enzyme-linked immunosorbent assay (ELISA). The messenger ribonucleic acid (mRNA) and protein expression levels of transforming growth factor-β1 (TGF-β1), as well as the bone morphogenetic protein 2 (BMP2) in the femoral head in each group, were determined using Reverse Transcription-Polymerase Chain Reaction (RT-PCR) and Western blotting. The rabbit model of SIONFH was successfully established. Compared with Control group, the Model group had a severer pathological injury of the femoral head, a lower level of VEGF in the femoral head, significantly decreased mRNA and protein levels of TGF-β1 and BMP2. Compared with Model group, the SR group had markedly improved pathological injury of the femoral head, a higher level of VEGF in the femoral head, significantly increased mRNA and protein levels of TGF-β1, as well as BMP2. SR can remarkably improve the pathological injury of the femoral head and increase the expression of VEGF in SIONFH rabbits, whose potential mechanism may be related to the activation of the TGF-β1/BMP2 signaling pathway.

Correlation of APOBEC3G expression with liver function indexes of patients with chronic hepatitis B and comparison in chronic hepatitis B, liver cirrhosis and liver cancer

Correlation of APOBEC3G expression with liver function indexes of patients with chronic hepatitis B and its expression in chronic hepatitis B, liver cirrhosis and liver cancer were investigated to evaluated the significance of APOBEC3G. Fifty-eight patients with chronic hepatitis B were selected, including 20 cases of chronic hepatitis B, 19 cases of liver cirrhosis and 19 cases of liver cancer. Liver function indexes were detected and analyzed, and messenger ribonucleic acid (mRNA) and protein expression levels of APOBEC3G in liver tissues were detected via reverse transcription-polymerase chain reaction (RT-PCR), western blotting and immunohistochemistry, followed by correlation analysis. Certain liver function indexes had significant differences among the three groups of patients (P<0.05). Results of RT-PCR, Western blotting and immunohistochemistry confirmed that the content of APOBEC3G in liver tissues was the highest in patients with chronic hepatitis B, slightly lower in patients with liver cirrhosis and the lowest in patients with liver cancer. The content of APOBEC3G mRNA in liver tissues had a certain correlation with the content of alanine aminotransferase (ALT) (r2 =0.34, P<0.05). Other liver function indexes had no significant correlations with APOBEC3G (P>0.05). APOBEC3G expression has a certain correlation with some liver function indexes of patients with chronic hepatitis B. There are significant differences in the expression level of APOBEC3G in patients with hepatitis, liver cirrhosis and liver cancer.

Hyperhomocysteinemia inhibits tibial fracture healing in rats through PI3K/AKT signaling pathway.

To explore the influence of hyperhomocysteinemia (hHcys) on the tibial fracture healing in rats and its effect on the phosphatidylinositol 3-hydroxy kinase (PI3K)/protein kinase B (AKT) signaling pathway. A total of 36 Sprague-Dawley rats were randomly divided into sham group (n=12), tibial fracture group (n=12) and hHcys + fracture group (n=12). The rats in tibial fracture group underwent the tibial fracture surgery, while the model of tibial fracture and hHcys was established in hHcys + fracture group. The level of plasma homocysteine (Hcy) in each group was analyzed using the full-automatic biochemical analyzer, the fracture stress biomechanical measurement was performed, and the ultimate bending strength and torque were calculated. Moreover, the protein expressions of PI3K and phosphorylated (p)-AKT in tibial tissues were detected using western blotting, the messenger ribonucleic acid (mRNA) levels of Bcl-2 associated X protein (Bax) and caspase-3 were detected using quantitative polymerase chain reaction (qPCR), the apoptosis was detected via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining, and the expressions of inflammatory factors were detected via immunohistochemistry. Compared with sham group, tibial fracture group and hHcys + fracture group had a significantly increased level of plasma Hcy, significantly decreased ultimate bending strength and torque, obviously decreased relative protein expressions of PI3K and p-AKT, increased mRNA levels of Bax and caspase-3 and an increased expression of pro-inflammatory factor tumor necrosis factor-α (TNF-α). Compared with tibial fracture group, hHcys + fracture group had a higher level of plasma Hcy, lower ultimate bending strength and torque, lower relative protein expressions of PI3K and p-AKT, higher mRNA levels of Bax and caspase-3, a higher apoptosis rate and a higher expression of TNF-α. hHcys blocks the downstream apoptotic signal transduction, promotes apoptosis and inflammatory response, and affects fracture healing through affecting the PI3K/AKT signaling pathway.

Expression and clinical significance of p62 protein in colon cancer.

p62 is a multifunctional protein involved in multiple cellular processes including proliferation, drug sensitivity and autophagy-associated cancer cell growth. However, the role of p62 in colon cancer remains controversial. Here we investigated the expression of p62 protein in colon cancer and its clinical significance.Patients with colon adenocarcinoma who underwent resection at the Third Affiliated Teaching Hospital of Xinjiang Medical University (Affiliated Cancer Hospital) were retrospectively analyzed. The expression of p62 protein in tumor tissues and adjacent normal tissues was detected by immunohistochemistry and western-blotting. Real-time quantitative polymerase chain reaction was used to detect the expression level of p62 messenger ribonucleic acid in specimens. Progression-free survival (PFS) and overall survival (OS) were assessed using Kaplan-Meier method and the log-rank test.A total of 85 colon cancer patients were enrolled, including 55 (64.71%) patients with high p62 expression, and 30 (35.29%) patients with low p62 expression. The transcription and expression level of p62 in colon cancer tissues were higher than those in adjacent normal tissues (P < .01). High expression of p62 was an independent risk factor for the poor prognosis (PFS and OS) of colon cancer.p62 may be a potential indicator of determining the progression and prognosis evaluation of colon cancer.

Long Noncoding RNA ENST00000581794.1 in Peripheral Blood Mononuclear Cells Acts as a Novel Biomarker in Patients with Acute Myocardial Infarction

The aim of this investigation is to investigate the long noncoding ribonucleic acid and mRNA expression profiles of acute myocardial infarction patients and matched healthy controls particularly the higher-ranked long noncoding ribonucleic acid ENST00000581794.1 in co-expression. This study group found that ENST00000581794.1 was up-regulated in acute myocardial infarction patients in pre-experiment. It is suggested that ENST00000581794.1 may be a potential biomarker of acute myocardial infarction. Fifty eight patients with acute myocardial infarction and 58 healthy controls were included in this study from September 2017 to February 2018. Venous blood samples were collected from patients within 6 h from the onset of chest pain and RT-qPCR measured the expression levels of long noncoding ribonucleic acid in peripheral blood mononuclear cells. All patients were followed for 12±1 mo. The expression levels of ENST00000581794.1 were significantly different between the two groups. The receiver operating characteristic curve showed that long noncoding ribonucleic acid ENST00000581794.1 (area under the curve = 0.744, 95 % confidence interval: 0.61~0.88, p=0.004) had a predictive effect on major adverse cardiovascular events. Survival curve analysis demonstrated that the incidence of major adverse cardiovascular events became significantly higher in patients with high expression of ENST00000581794.1 as the observation time increased. Multivariate regression analysis indicated that ENST00000581794.1 was an independent predictor of major adverse cardiovascular events in patients with acute myocardial infarction (hazard ratio: 1.17, 95 % confidence interval: 1.00~1.37, p=0.048). Highly expressed ENST00000581794.1 in peripheral blood mononuclear cells may be used as a biomarker for diagnosis and prognosis of acute myocardial infarction.

Infection of Placental Extravillous Cytotrophoblasts with Human Cytomegalovirus Causes a Treg/Th17 Imbalance at the Maternal–Fetal Interface

This paper aimed to evaluate whether human cytomegalovirus (HCMV) infection in extravillous cytotrophoblasts (EVT) could shift the balance between regulatory T (Treg) and T-helper type 17 (Th17) cells in vitro. In this study, primary EVT isolated from first trimester placental tissues were infected with HCMV, and conditional media were harvested after cultivation for 72 h. T lymphocytes were cultured in the presence or absence of HCMV-infected conditional media. The frequencies of Th17 or Treg cells from HCMV group were significantly lower or higher than those from the control group, with the expression of corresponding key cytokines at both messenger ribonucleic acid and secretion levels, respectively. The ratio of Treg to Th17 cells was significantly lower in HCMV group than that in control group (P < 0.01). In conclusion, tiled Th17/Treg balance at maternal–fetal interface exists after HCMV infection.

Glutamine protects myocardial ischemia-reperfusion injury in rats through the PI3K/Akt signaling pathway.

To study the protective mechanism of glutamine (Gln) on myocardial ischemia-reperfusion (IR) injury in rats through the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway. A total of 30 healthy SD rats weighing 200-300 g were used in this experiment. They were randomly divided into 3 groups: sham group (n=10), myocardial IR injury group (IR group, n=10), IR+Gln group (n=10). The protein expression levels of phosphorylated Akt (p-Akt), total Akt (t-Akt), phosphorylated mammalian target of rapamycin (p-mTOR), mTOR, proliferating cell nuclear antigen (PCNA), P21, and Tubulin were determined by Western blotting (WB). Quantitative Polymerase Chain Reaction (qPCR) was applied to detect the messenger ribonucleic acid (mRNA) levels of Akt and mTOR. 3-(4,5)-dimethylthiazol(-z-y1)-3,5-diphenyl tetrazolium bromide (MTT) test was utilized to examine the proliferation ability of cardiomyocytes in vitro. Besides, the contents of the inflammatory cytokines, tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6) were measured via enzyme-linked immunosorbent assay (ELISA). Cell apoptosis in each group was examined through Hoechst staining. Compared with those in the sham group, ratios of p-AKT/AKT, p-mTOR/mTOR, and the level of PCNA extremely significantly decreased, but the level of P21 notably increased in IR group (p<0.01). In comparison with those in the IR group, ratios of p-AKT/AKT, p-mTOR/mTOR, and the level of PCNA were remarkably raised, while the level of P21 was remarkably reduced in IR+Gln group (p<0.05). QRT-PCR results manifested that there were no significant differences in the mRNA levels of Akt and mTOR among the three groups [no significant difference (NS)]. Moreover, the cell proliferation ability in IR group was remarkably lower than that in the sham group (p<0.01), while it was enhanced in the IR+Gln group compared with that in the IR group (p<0.05). Additionally, IR group had significantly elevated expression levels of TNF-α and IL-6 compared with the sham group (p<0.01), whereas the IR+Gln group had notably decreased expression levels of TNF-α and IL-6 compared with IR group (p<0.05). In comparison with that in the sham group, the apoptosis in IR group was significantly raised (p<0.01), and compared with that in the IR group, the apoptosis in the IR+Gln group prominently decreased (p<0.05). The contents of the inflammatory cytokines, TNF-α, and IL-6 presented the same trends among the three groups. Gln activates the PI3K/Akt signaling pathway by increasing the levels of p-AKT and p-mTOR. Gln can increase the PCNA level and reduce the P21 level, so as to enhance the proliferation ability of cardiomyocytes. Besides, Gln reduces the levels of inflammatory cytokines, TNF-α, and IL-6, and inhibits cell apoptosis. Finally, Gln can protect cells from myocardial IR injury by activating the PI3K/Akt signaling pathway.

Simvastatin promotes osteogenic differentiation of mesenchymal stem cells in rat model of osteoporosis through BMP-2/Smads signaling pathway.

By establishing osteoporosis (OP) model in rats, the specific regulatory effect of simvastatin on promoting the differentiation of mesenchymal stem cells (MSCs) into osteoblasts through the bone morphogenetic protein 2 (BMP-2)/Smads signaling pathway was investigated. A total of 45 Sprague-Dawley rats were selected to establish the OP model by performing ovariectomy. The rats were divided into OP model group (OP group, n=15), 10-7 mmol/L simvastatin treatment group (SIM group, n=15), and normal control group (Control group, n=15). After the experimental period, the enzyme-linked immunosorbent assay (ELISA) was applied to observe the serum levels of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and IL-1. Reverse Transcription-Polymerase Chain Reaction (RT-PCR) was adopted to detect the contents of the differentiation-associated genes [runt-related transcription factor 2 (RUNX2) and Osterix (Osx)]. Later, the bone marrow MSCs (BMSCs) were selected and divided into Control group, 10-7 mol/L simvastatin group (SIM group), and osteoinduction medium group (OM group). Cell morphology in each group was observed. The Cell Counting Kit-8 (CCK-8) was performed to determine the proliferation activity of BMSCs. ELISA was performed to measure the level of alkaline phosphatase (ALP). RT-PCR was conducted to examine the levels of key differentiation-associated gene RUNX2 and those in BMP-2/Smads pathway. Moreover, the Western blotting was adopted to analyze the expressions of RUNX2 and genes in BMP-2/Smads pathway. The serum levels of TNF-α, IL-6, and IL-1 in OP group were remarkably higher than those in the Control group, and their levels in the SIM group were close to those in the Control group. The elevated messenger ribonucleic acid (mRNA) levels of the key differentiation-associated factors RUNX2, osteoprotegerin (OPG), osteopontin (OPN), and Osx were observed in the SIM group. In vitro cell culture revealed that the cells were in a favorable growth status in the SIM group and OM group, mostly manifesting in fusiform or spindle shape, and proliferated rapidly. In addition, the ALP level notably increased in the two groups compared with that in the Control group (p<0.05). Both SIM group and OM group had evidently higher mRNA expression levels of RUNX2, OPG, OPN, and Osx than those in the Control group (p<0.05), consistent with the expression trends of the genes in BMP-2/Smads pathway. The Western blotting indicated that the expression levels of RUNX2 and genes in BMP-2/Smads pathway in the SIM group were significantly higher than those in the Control group. Simvastatin can promote the differentiation of MSCs into osteoblasts in the OP rat model through the BMP-2/Smads signaling pathway.

Relationship between NogoA/NgR1/RhoA signaling pathway and the apoptosis of cerebral neurons after cerebral infarction in rats.

The aim of this study was to investigate the effect and the mechanism of the NogoA/NgR1/RhoA signaling pathway on the apoptosis of neurons in cerebral infarction (CI) rats. Our findings might provide references for clinical prevention and treatment of CI. A total of 60 adult male Wistar rats were randomly divided into 3 groups, including: Sham operation group (Sham group), CI group, and CI + NogoA gene knockout group (CI + NogoA KO group) using a random number table. The model of CI was successfully constructed using suture method in rats of CI group and CI + NogoA KO group. Only blood vessels were exposed in Sham group. At 2 days after CI operation, the rats were killed, and brain tissues were collected. Reverse Transcription-Polymerase Chain Reaction (RT-PCR) and Western blotting were used to detect the messenger ribonucleic acid (mRNA) and protein expression levels of NogoA/NgR1/RhoA in brain lesion tissues of rats in the three groups, respectively. Subsequently, the pathological damage of brain tissues was detected via hematoxylin and eosin (H&E) staining. TTC staining was carried out to evaluate the infarction area in each group. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining was conducted to measure the apoptosis level of neurons in brain tissues of rats in each group. Additionally, the level of Nissl's body in brain tissues of each group was examined by Nissl staining. Furthermore, the expression level of the platelet-derived growth factor (PDGF) in brain tissues of rats in the three groups was measured via immunohistochemistry. The mRNA and protein expression levels of the NogoA/NgR1/RhoA signaling pathway in brain tissues of rats in CI group increased significantly (p<0.05). NogoA KO could significantly reduce the infarction area of brain tissues in rats (p<0.05). H&E staining and Nissl's body staining revealed that neurons in the brain tissues of rats showed evident edema and disordered arrangement after CI. Meanwhile, the number of Nissl's body was remarkably reduced. However, after KO of NogoA, brain tissue damage was significantly alleviated in rats, and the number of Nissl's body increased remarkably at the same time (p<0.05). According to TUNEL staining results, inhibiting NogoA could notably reverse CI-induced apoptosis of neurons in brain tissues of rats (p<0.05). Immunohistochemical staining results demonstrated that the expression of PDGF in brain tissues of rats in CI group decreased markedly, whereas was significantly elevated in rats of CI + NogoA KO group (p<0.05). The expression of the NogoA/NgR1/RhoA signaling pathway was significantly elevated in brain tissues of CI rats. Furthermore, suppressing the NogoA/NgR1/RhoA signaling pathway could reduce CI-induced apoptosis of neurons in rats.

Effect of rosiglitazone on myocardial injury in septic rats through NF-κB pathway.

To explore the effect of rosiglitazone on myocardial injury in septic rats through the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway. A total of 60 healthy adult female Sprague-Dawley (SD) rats were randomly divided into 4 groups, namely, group A (sepsis model group, n=15), group B (sham operation group, n=15), group C (sepsis model + 3 mg/kg rosiglitazone, n=15), and group D (sham operation group + 3 mg/kg rosiglitazone, n=15), respectively. After the sepsis model was successfully established, the rats were administered with 3 mg/kg rosiglitazone by gavage, with a gavage volume of 1 mL, once a day for a total of 3 days. Blood was taken from the abdominal aorta, while lactate dehydrogenase (LDH) and creatine phosphokinase kits were used to detect the levels of LDH and creatine phosphokinase in serum. Then, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining was adopted to identify myocardial tissue apoptosis, hematoxylin and eosin staining (H&E) was applied to detect myocardial tissue morphology, and enzyme-linked immunosorbent assay (ELISA) was utilized to examine the protein expression level of tumor necrosis factor-alpha (TNF-α) in rat serum. Subsequently, the messenger ribonucleic acid (mRNA) level of TNF-α in myocardial tissues was measured via fluorescence quantitative Reverse Transcription-Polymerase Chain Reaction (qRT-PCR) method, and the activity of NF-κB was detected by electrophoretic mobility shift assay (EMSA). Compared with those in group A, apoptotic cells in group B and group D were notably increased (p<0.05). At 3 days after administration with rosiglitazone (3 mg/kg), apoptotic cells were markedly decreased (p<0.05). H&E staining results manifested that 3 mg/kg rosiglitazone prominently improved myocardial tissue morphology in rats. The protein level of TNF-α in serum, the mRNA expression level of TNF-α in myocardial tissues, and the activity of NF-κB in group C treated with rosiglitazone were lower than those in group A (p<0.05). Rosiglitazone can alleviate myocardial injury in septic rats by suppressing the TNF-α expression and this process is associated with the regulation on the NF-κB signal transduction pathway.

FOXN3 Inhibits Cell Proliferation though Regulating E2F1 in Prostate Cancer

In this study, it was demonstrated that forkhead box N3 was downregulated in prostate cancer tissues compared to the matched normal adjacent tissues. Prostate cancer patients with high forkhead box N3 expression more frequently had longer survival (P<0.05) than those with low forkhead box N3 expression. Forkhead box N3 inhibited prostate cancer cell proliferation in vitro and in vivo. In addition, forkhead box N3 repressed expression of E2F1, a reported potential oncogene at the messenger ribonucleic acid and protein levels. Taken together, these data delineate an unrecognized function of forkhead box N3 as a tumor suppressor in prostate cancer via repressing the expression of E2F1.

Enhanced UV Resistance Role of Death Domain–Associated Protein in Human MDA-MB-231 Breast Cancer Cells by Regulation of G2 DNA Damage Checkpoint

Purpose: Death domain–associated protein (DAXX) is a multifunctional nuclear protein involved in apoptosis, transcription, deoxyribonucleic acid damage response, and tumorigenesis. However, the role of DAXX in breast cancer development and progression remains elusive. In this study, we examined the expression patterns and function of DAXX in human breast cancer samples and cell lines. Methods: Immunohistochemistry was used to analyze the expression and localization patterns of DAXX. Additionally, we investigated whether DAXX played an intrinsic role in the cellular response to damage induced by ultraviolet (UV) irradiation in MDA-MB-231 breast cancer cells (isolated at M D Anderson from a pleural effusion of a patient with invasive ductal carcinoma). Results: Our results showed that nucleus size, chromatin organization, and DAXX localization were altered in breast cancer tissues compared with those in control tissues. Compared with cytoplasmic and nuclear expression in benign breast tissues, DAXX was colocalized with promyelocytic leukemia in nuclei with a granular distribution. Endogenous DAXX messenger ribonucleic acid levels were upregulated upon UV radiation in MDA-MB-231 cells. DAXX-deficient cells tended to be more sensitive to irradiation than control cells. Conversely, DAXX-overexpressing cells exhibited reduced phosphorylated histone H2AX (γ-H2AX) accumulation, increased cell survival, and resistance to UV-induced damage. The protective effects of DAXX may be related to the activation of the ataxia telangiectasia mutated (ATM)-checkpoint kinase 2 (ATM-CHK2)-cell division cycle 25c (CDC25c) signaling pathways in Gap2/Mitosis (G2/M) checkpoint and ultimately cell cycle arrest at G2/M phase. Conclusions: Taken together, these results suggested that DAXX may be an essential component in breast cancer initiation, malignant progression, and radioresistance.

Cerebral toxoplasmosis in HIV-infected patients over 2015-2018 (a case study of Russia).

Cerebral toxoplasmosis is a leading cause of the central nervous system disorders in acquired immune deficiency syndrome. This study aimed to investigate the clinical course of cerebral toxoplasmosis in human immunodeficiency virus (HIV)-infected individuals. The study included 90 HIV-infected patients with cerebral toxoplasmosis, who underwent inpatient treatment. In case of positive enzyme immunoassay, HIV infection was confirmed with the immunoblot test. The HIV-1 ribonucleic acid level was determined using the polymerase chain reaction method. The flow cytometry was used for counting CD4 (cluster of differentiation 4 cells). Pathomorphological examination included the autopsy, gross and microscopic examination of internal organs, histological and other methods. The incidence of cerebral toxoplasmosis significantly increases at the CD4 count below 100 cells/μl, P < 0.001, and at the HIV viral load above 50 copies/ml, P < 0.05. The clinical picture of cerebral toxoplasmosis included focal symptoms, cognitive impairment, toxic syndrome, mild cerebral symptoms and a meningeal symptom. Given the absence of a specific clinical picture and the absence of abnormal laboratory and instrumental findings, the cerebral toxoplasmosis needs to be diagnosed with a number diagnostic methods combined: clinical examination, laboratory testing, immunological examination, molecular genetic testing and neuroradiological imaging.