Tumor Targeting via siRNA-COG3 to Suppress Tumor Progression in Mice and Inhibit Cancer Metastasis and Angiogenesis in Ovarian Cancer Cell Lines.

BACKGROUND: The COG complex is implicated in the tethering of retrograde intra-Golgi vesicles, which involves vesicular tethering and SNAREs. SNARE complexes mediate the inva-sion and metastasis of cancer cells through MMPs which activate growth factors for ECM frag-ments by binding to integrin receptors. Increasing MMPs is in line with YKL40 since YKL40 is linked to promoting angiogenesis through VEGF and can increase ovarian cancer (OC) resistance to chemotropic and cell migration. OBJECTIVE: The aim of this study is an assessment of siRNA-COG3 on proliferation, invasion, and apoptosis of OC cells. In addition, siRNA-COG3 may prevent the growth of OC cancer in mice with tumors. METHODS: Primary OC cell lines will be treated with siRNA-COG3 to assay YKL40 and identified angiogenesis by Tube-like structure formation in HOMECs. The Golgi morphology was analyzed using Immunofluorescence microscopy. Furthermore, the effects of siRNA-COG3 on the prolifer-ation and apoptosis of cells were evaluated using MTT and TUNEL assays. Clones of the HOSEpiC OC cell line were subcutaneously implanted in FVB/N mice. Mice were treated after two weeks of injection of cells using siRNA-COG3. Tumor development suppression was detected by D-luciferin. RT-PCR and western blotting analyses were applied to determine COG3, MT1-MMP, SNAP23, and YKL40 expression to investigate the effects of COG3 gene knockdown. RESULTS: siRNA-COG3 exhibited a substantial effect in suppressing tumor growth in mice. It dra-matically reduced OC cell proliferation and triggered apoptosis (all p < 0.01). Inhibition of COG3, YKL-40, and MT1-MPP led to suppression of angiogenesis and reduction of microvessel density through SNAP23 in OC cells. CONCLUSION: Overall, by knockdown of the COG3 gene, MT1-MMP and YKL40 were dropped, leading to suppressed angiogenesis along with decreasing migration and proliferation. SiRNA-COG3 may be an ideal agent to consider for clinical trial assessment therapy for OC, especially when an antiangiogenic SNAR-pathway targeting drug.

Role of S100 and YKL40 on Intraventricular Cerebral Hemorrhages in the Preterm Infant and the Neuroprotective Role of miR-138-siRNAs-HIF-1a and miR-21-siRNAs-HVCN1 in Neonatal Mice with Nerve Injury.

BACKGROUND: Epilepsy and intraventricular-cerebral hemorrhage is a common complication irreversible in preterm infants. Inflammation leads to an increase in intracellular calcium, acidosis, and oxygen usage, and finally, may damage brain cells. Increases in HIF-1a and HVCN1 can reduce the complications of oxygen consumption and acidosis in infants with intraventricular hemorrhage (IVH). On the other hand, decreases in S100B can shield nerve cells from apoptosis and epilepsy by reducing brain damage. OBJECTIVE: In this research, we investigated how miR-138-siRNAs-HIF-1a and miR-21-siRNAs-HVCN1 affect apoptosis in hypoxic mice. METHODS: On the first and third days after delivery, the YKL40, HIF-1a, HVCN1, and S100b genes were compared between two groups of preterm infants with and without maternal inflammation. Afterward, the miRNAs were transfected into cell lines to monitor variations in YKL40, HIF-1a, HVCN1, and S100b gene expression and nerve cell apoptosis. We changed the expression of S100b, HVCN1, and HIF-1a genes by using specific siRNAs injected into mice. Using real-time PCR, Western blotting, flow cytometry (FCM), and immunofluorescence, and changes in gene expression were evaluated (IHC). RESULTS: HVCN1 gene expression showed a strong negative correlation with epilepsy in both groups of infants (P<0.001). Significant correlations between epilepsy and the expression levels of the S100b, YKL40, and HIF-1a genes were found (P<0.001). According to FCM, after transfecting miRNA-431 and miRNA-34a into cell lines, the apoptosis index (A.I.) were 41.6 3.3 and 34.5 5.2%, respectively, while the A.I. were 9.6 2.7 and 7.1 4.2% after transfecting miRNA-21 and miRNA-138. MiR-138-siRNAs-HIF-1a and miR-21-siRNAs-HVCN1 were simultaneously injected into hypoxic mice, and IHC double-labeling revealed that this reduced apoptosis and seizures compared to the hypoxic group. CONCLUSION: Our findings demonstrate that miR-138-siRNAs-HIF-1a and miR-21-siRNAs-HVCN1 injections prevent cerebral ischemia-induced brain damage in hypoxia mice by increasing HVCN1 and HIF-1a and decreasing S100b, which in turn lessens apoptosis and epilepsy in hypoxic mice.

The Shift of HbF to HbA under Influence of SKA2 Gene; A Possible Link between Cortisol and Hematopoietic Maturation in Term and Preterm Newborns.

BACKGROUND: We hypothesized that the SKA2 gene can convert hemoglobin F to A leading to the maturity of the hematopoietic system by glucocorticoid hormone; so, the present study aimed to investigate the health outcome of newborns by using the effect of SKA2 gene on hematopoietic maturation. METHODS: At first, 142 samples were divided into term and preterm. After sampling from the umbilical cord blood, the expression of SKA2 genes and HbA and F were evaluated by quantitative RT-PCR. The blood gases were measured by Campact 3 device. Finally, the cortisol level was measured by ELISA method and HbA and F levels were investigated by capillary electrophoresis. RESULTS: The blood gases and Apgar scores were more favorable in term newborns (P <0.001). Levels of protein/expression of HbF in newborns with Apgar score greater than 7 was lower than that of the newborns with Apgar score below 7 (P <0.001). Cortisol and HbA levels were considerably higher in term newborns compared to the preterm ones (P <0.001). In the preterm and term groups, SKA2 gene expression had a positive and significant relationship with cortisol and HbA levels as well as a negative relationship with the HbF level. In the preterm group, a positive and significant relationship was observed between the expression of SKA2 and HbF genes. CONCLUSION: The results revealed that the SKA2 gene affected hematopoietic maturation in preterm and term newborns and the health outcome of newborns improved by increasing HbA level.

The Correlation of SKA2 with Cortisol, IL-1ß and Anxiety in Pregnant Women with the Risk of Preterm Delivery.

OBJECTIVE: The association between preterm birth (PTB), Spindle and Kinetochore Associated Complex Subunit 2 gene (SKA2), cortisol and anxiety have been shown, but in this study, we aimed to clarify whether the expression of the SKA2 gene plays a role in interleukin1ß (IL-1ß) level since increasing level of IL-1ß is linked with PTB. METHODS: The case-control study was conducted on 49 and 51 women with preterm and term delivery, respectively. The score of anxiety was ranked according to the Spielberger state trait Anxiety Inventory. The concentration of cortisol and IL-1ß was determined by the ELISA method. The expression of SKA2 gene was assessed by the quantitative real time real time polymerase chain reaction (qRT-PCR). The western blot analysis was also performed to confirm the expression of SKA2 at the levels of protein. RESULTS: The results showed that the gene/protein expression of SKA2, the concentrations of cortisol and IL-1ß were significantly higher in the preterm than the term group. In the preterm group, the expression of SKA2 was positively correlated to the other factors including cortisol, IL-1ß, and the degree of anxiety. CONCLUSION: Our findings suggest that the expression of SKA2 was correlated positively to the levels of cortisol, IL-1ß and the rate of anxiety in women with PTB.

Association of GRP78, HIF-1α and BAG3 Expression with the Severity of Chronic Lymphocytic Leukemia.

INTRODUCTION: Parallel with the progression of Chronic Lymphocytic Leukemia (CLL), the levels of 78KDa Glucose-Regulated Protein (GRP78) and Hypoxia-Inducible Factor 1 alpha (HIF-1α) are increased as they may activate the induction of anti-apoptotic proteins such as BCL2 Associated Athanogene 3 (BAG3). Previous studies have indicated that there is a positive correlation among GRP78, HIF-1α and BAG3. OBJECTIVE: This study aimed to evaluate the effect of metabolic factors involved in invasive CLL on apoptotic factors. METHODS: A case-control study was conducted on 77 patients diagnosed with CLL along with 100 healthy individuals. Cell blood count was performed for all participants. According to Binet's classification, CLL patients were divided into different groups. B cells were isolated from the peripheral blood of CLL patients by binding to anti-CD19 beads. The expression of BAG3, GRP78 and HIF-1α genes was analyzed using the RT-PCR method. To confirm the results of RT-PCR, western blot analysis was carried out. RESULTS: The results showed that there was a strong association among the expression of BAG3, GRP78 and HIF-1α. The stage of CLL in patients was highly correlated with the expression rate of each gene (p<0.001). Accordingly, the western blot analysis indicated that the concentrations of GRP78 and HIF-1α were significantly higher than the expression of BAG3, considering the stage of CLL. CONCLUSION: It was shown that increased expression of GRP78 and HIF-1α could result in the elevation of BAG3, as well as the disease progression. Therefore, the role of these metabolic factors might be more pronounced compared with the anti-apoptotic agents to monitor disease progression in CLL patients.

SKA2 gene - A novel biomarker for latent anxiety and preterm birth prediction.

BACKGROUND: There is a relationship between preterm birth (PTB) and anxiety. Spindle and Kinetochore Associated Complex Subunit 2 (SKA2) gene polymorphism (NC_000017.11: g.59110368 G > A) has also been associated with the development of anxiety. The current study was designed to evaluate the relationship between SKA2 gene SNP (NC_000017.11: g.59110368 G > A) with the occurrence of anxiety and PTB which might be considered a predictive biomarker for the prediction of preterm delivery. METHODS: SKA2 gene (SNP rs7208505) genotyping was performed in 300 women with term birth (TB) and 293 women with PTB using PCR-RFLP method and then followed by DNA sequencing. Cortisol level was analyzed with ELISA method and the presence of anxiety was detected using Spielberg Inventory. RESULTS: The AA genotype of SKA2 gene significantly increased the risk of PTB compared to the GG genotype by 9.6 fold ([CI] 4.5-20.2, P < 0.001) according to codominant model. Also, the frequency of A allele was significantly higher in PTB group (χ2 = 20.4, df = 1, P < 0.001) in comparison with the control group that increased the risk of PTB by 1.703 fold ([CI] 1.39-2.23, P < 0.001). Women with higher cortisol level with average 343.7 ± 3 nmol/L had AA genotype, while, the concentrations of cortisol in women with AG, and GG genotypes were 244.2 ± 3.1 nmol/L and 192.6 ± 2.5 nmol/L, respectively (P < 0.001). The score of apparent and latent anxiety in women with the AA genotype was higher compared to the AG and GG genotypes and also this score in women with the AG genotype was higher than the GG genotypes (P < 0.001). The history of preterm delivery was higher in women with the AA genotype (42.1%) in comparison with the GG (14.9%) and AG (22%) genotypes (P < 0.05). CONCLUSION: The results of the current study suggest that prognosis of women with the AA genotype are more susceptible to be spontaneous preterm birth. Therefore, the A allele of SKA2 gene (NC_000017.11:g.59110368 G > A) could be as a predictive biomarker for the risk of PTB.