[Upregulation of miR-498 suppresses Th17 cell differentiation by targeting STAT3 in rheumatoid arthritis patients].

To investigate the effect and mechanism of miR-498 on Th17 cell differentiation of peripheral blood mononuclear cells (PMBCs) in rheumatoid arthritis (RA) patients, peripheral blood samples were collected from RA patients and healthy controls, respectively. The proportion of CD4+IL-17+ T cells (Th17 cells) or CD4+FOXP3+ T cells (Tregs) in T cells and the Th17/Treg ratio were identified by the flow cytometer. The STAT3 and miR-498 expression were measured by Western blot and real-time PCR, respectively. ELISA was used to detect IL-17 concentrations. Luciferase assay was performed to confirm that miR-498 directly targeted the 3' untranslated region (3'UTR) of STAT3 in CD4+ T cells. The effect of miR-498 on Th17 cell differentiation was explored by transfection of miR-498 mimic and/or pcDNA-STAT3 into CD4+ T cells. In PMBCs of RA patients, the Th17/CD4+ T cell ratio was significantly increased, while the Tregs/CD4+ T cell ratio was obviously decreased, leading to a higher Th17/Treg ratio. The results showed a reduced miR-498 expression and an increased STAT3 protein expression in PMBCs, and an increased IL-17 concentration in serum of RA patients. In cells transfected with wild-type-STAT3-LU, miR-498 mimic significantly reduced the luciferase activity, STAT3 gene and protein expression, and miR-498 inhibitor had an opposite function. While the miR-498 mimic/inhibitor had no effect on the luciferase activity and STAT3 expression in cells transfected with mutant-STAT3-LU. CD4+ T cells transfected with miR-498 mimic had a lower Th17/CD4+ T cell ratio and IL-17 concentration, however, transfection of pcDNA-STAT3 reversed the effect of miR-498 mimic on Th17/CD4+ T cell ratio and IL-17 concentration. These results suggest that overexpression of miR-498 suppresses Th17 cell differentiation by targeting STAT3 in RA patients.

Sialic Acid and Iron Content in Breastmilk of Chinese Lactating Women.

OBJECTIVE: To study sialic acid and iron content in breastmilk in Chinese women during different lactation stages. METHODS: Sialic acid and iron content of colostrum, transitional milk, mature milk, and involutional milk were determined using a neuraminidase assay kit and the ferrozine method, respectively in 88 lactating women (58 Term, 30 Preterm). RESULTS: The mean (SD) sialic acid levels of colostrum, transitional milk, mature milk, and involutional milk were 2201.4 (676.6) mg/L, 1445.9 (423.4) mg/L, 395.3 (96.0) mg/L and 273.0 (76.9) mg/L, respectively. The median iron content were 0.05 mg/L, 0.06 mg/L, 0.25 mg/L and 0.35 mg/L, respectively, in successive stages of lactation. Sialic acid and iron were significantly higher in breast milk of preterm mothers compared to term mothers. CONCLUSION: Sialic acid and iron content in breast milk vary greatly throughout the lactation stages, which probably reflects the infants' needs for growth and development at different stages.

Serum Concentrations of Antibodies against Outer Membrane Protein P6, Protein D, and T- and B-Cell Combined Antigenic Epitopes of Nontypeable Haemophilus influenzae in Children and Adults of Different Ages.

Nontypeable Haemophilus influenzae (NTHi) is one of the most common etiologies of acute otitis media, rhinosinusitis, and pneumonia. Outer membrane proteins (OMPs) are the main focus in new vaccine development against NTHi, as the H. influenzae type b (Hib) vaccine does not cover noncapsulated NTHi. The OMPs P6 and protein D are the most promising candidate antigens for an NTHi vaccine, and low antibody levels against them in serum may be correlated with infection caused by NTHi. In the current study, we measured the antibody titers against P6, protein D, and their T- and B-cell combined peptide epitopes in healthy individuals of different ages. We found that children <1 month old had the lowest antibody levels against NTHi P6, protein D, and their T- and B-cell combined antigenic epitopes. Antibody titers increased at ages 1 to 6 months, peaked at 7 months to 3 years, and remained high at 4 to 6 years. The antibody titers started to decrease after 6 years and were the lowest in the 21- to 30-year group. The geometric mean titers (GMTs) of T- and B-cell combined antigenic epitopes in P6 and protein D were positively correlated with those of the protein antigens. Among 12 peptides tested, P6-61, P6-123, and protein D-167 epitopes were better recognized than others in human serum. These findings might contribute to the development of an effective serotype-independent vaccine for H. influenzae.

Insulin promotes proliferation and migration of breast cancer cells through the extracellular regulated kinase pathway.

The present study was undertaken to determine the roles of insulin in the growth of transplanted breast cancer in nude mice, and the proliferation and migration of MCF-7 human breast cancer cells and assess its influence on downstream signaling pathways. In a xenograft mouse model with injection of MCF-7 human breast cancer cells, tumor size was measured every other day. The insulin level and insulin receptor (IR) were increased in the breast cancer patient tissues. Insulin injected subcutaneously around the tumor site in mice caused increase in the size and weight of tumor masses, and promoted proliferation and migration of MCF-7 cells. The effects of insulin on the increase in the proliferation and migration of MCF-7 human breast cancer cells were abolished by pretreatment with the extracellular regulated kinase (ERK) inhibitor PD98059. Insulin increased the phosphorylation of ERK in the MCF-7 cells. These results indicate that insulin promotes the growth of breast cancer in nude mice, and increases the proliferation and migration of MCF-7 human breast cancer cells via the ERK pathway.

[Effect of miR-15a on induction of apoptosis in breast cancer MCF-7 cells].

OBJECTIVE: To investigate the effect and mechanism of miR-15a on the induction of apoptosis in breast cancer cells. METHODS: To detect the expression level of miR-15a in breast cancer cell line MCF-7 cells and human mammary gland epithelial cell line MCF-10A cells by quantitative PCR. The target point of MCF-7 was predicted by software and was validated by luciferase report gene system. MiR-15a was transfected into MCF-7 cells with liposomes. The expression of Bcl-2 in MCF-7 cells was detected by Western blotting and the apoptosis rate of MCF-7 cells was detected by flow cytometry. RESULTS: The expression level of miR-15a in MCF-7 cells was lower than that in the MCF-10A cells (0.253:1, P < 0.0001). The expression of MiR-15a was significantly inhibited by Bcl-2 (P < 0.05). Compared with the control, Bcl-2 expression was significantly decreased in the MCF-7 cells. The results of flow cytometry showed that the apoptosis rate was 13.4% in non-transfected MCF-7 cells, 15.9% in MCF-7 cells transfected with control RNA, and 31.5% in MCF-7 cells transfected with miR-15a (P < 0.05), indicating an evident induction of apoptosis in the MCF-7 cells. CONCLUSION: miR-15a may have a potential application value in breast carcinoma biotherapy.