JAK2 Mediates the Regulation of Pept1 Expression by Leptin in the Grass Carp (Ctenopharyngodon idella) Intestine.

Oligopeptide transporter 1 (Pept1) is located on the brush border membrane of the intestinal epithelium and plays an important role in dipeptide and tripeptide absorption from protein digestion. In this study, we cloned and characterized the cDNA sequence of Janus kinase 2 (JAK2) from Ctenopharyngodon idella. The expression patterns of JAK2 in various tissues and developmental stages were characterized by quantitative real-time PCR (qRT-PCR). The mRNA expression levels of JAK2 and Pept1 regulated by leptin in the intestine were also analyzed in vitro and in vivo. The cDNA sequence of JAK2 is 3378 bp in length, and the mRNA of JAK2 was broadly expressed in all tissues and embryonic stages of C. idella analyzed. In addition, we found that leptin regulated expression of JAK2 and Pept1 in the intestine; Pept1 expression was down-regulated by the JAK2 inhibitor AG490 in vivo and in vitro. Furthermore, luciferase experiments showed that overexpression of the JAK2 gene significantly upregulated the activity of the Pept1 5' regulatory sequence in C. idella. In conclusion, these results may help in elucidating the regulatory effect of the leptin-mediated JAK2 pathway on intestinal Pept1 expression in C. idella and the molecular mechanism of peptide transport by the intestinal transporter Pept1 in fishes.

[Evaluation and Comparison of Thromboelastography and Conventional Coagulation Tests for Blood Coagulation Function in Children with Henoch-Schönlein Purpura].

OBJECTIVE: To investigate the efficacy of disease control, survival time and safely in treatment of newly diagnosed multiple mycloma patients with different dose of tenalidomide regimens. METHODS: The clinical data of 116 patients with multiple myeloma from June 2011 to June 2015 were collected and analyzed retrospectively. According to doses of used lenalidomide based on dexamethasone plus lenalidomide regimen 116 patients were divided into 2 groups: conventional dose group (58 cases) and low dose group (58 cases). The ORR, PFS rate and OS rate during followed-up for 3 years, KPS score, RNS score and immunophenotypic index before and after treatment and drug toxicity incidence were compared between 2 groups. RESULTS: The ORR for 2 treatment courses of low dose group was significantly lower than that in conventienal dose group (P＜0.05). The ORR for 4 and 6 treatment courses was not significantly different between 2 groups (P＞0.05). The PFS rate and OS rate during followed-up for 3 years was no significantly different between 2 groups (P＞0.05). The KPS score and RNS score after treatment of low dose group were significantly better than those in conventional dose group and before treatment (P＜0.05). The levels of immunophenotypic index after treatment of both groups were significantly better than those before treatment (P＜0.05). The incidence of III-IV grade hematological toxicity, pulmonary infection and herpes were not significantly different between 2 groups (P＞0.05). The incidence of peripheral neuropathy and gastrointestinal reactions in the low dose group were significantly lower than that in conventional dose group (P＜0.05). CONCLUSION: Conventional and low doses of lenalidomide possess the same control effects and survival time for treatment of newly dingnosed patients with multiple myeloma; Despite, the initiation of effects from the low dose lenalidomide is relatively slower, it contributes to raise the overall quality of life and reduce the risk of drug toxicity.

[Levels and Significance of Serum Interleukin-35 and the New Regulatory T Cells -iTR35 Cells in Myelodysplastic Syndrome Patients].

OBJECTIVE: To investigate the significance of serum interleukin-35 level and the new regulatory T cells -iTR35 cells in patients with myelodysplastic syndrome (MDS). METHODS: Twenty three cases of newly diagnosed MDS were enrolled in this study from January 2014 to January 2016 in Department of Hematology of The First Hospital of Quanzhou in Fujian Province. According to MDS International Prognostic Scoring System (IPSS), the 23 patients were divided into 4 groups: high-risk (n=4), intermediate risk-2 (n=10), intermediate risk-1 (n=5) and low-risk group(n=4). Twenty healthy people of routine physical examination were used as control during the same period. Enzyme Linked Immunosorbent assay(ELISA) and flow cytometry(FCM) were used to detect the expression level of serum IL-35, the proportion of iTR35 cells, and the expression levels of associated molecules such as IL-12p35,IL-27EBl3 respectively. RESULTS: The proportion of CD4+ CD25+ Foxp3+ Treg cells in peripheral blood of MDS patients was significantly higher than that in controls (P<0.01). The proportion of iTR35 cells was also higher than that in controls(P<0.01). However, the proportion of CD4+ CD25- Foxp3+ T cells was not significantly different between 2 groups (P>0.05). The level of serum IL-35 in MDS patients was significantly higher than that in control group (P<0.05). The expression levels of IL-12p35 and IL-27EBl3 in the Treg cells were also significantly upregulated than those in control group(P<0.05), the expression levels of IL-35, IL-12p35 and IL-27EBl3 in MDS group positively correlated with the proportion of iTR35 cells(r=0.92, 0.99 and 0.52, P<0.05, respectively). IL-35 level and the proportion of iTR35 cells in 4 groups of MDS showed significantly difference in general term, no significant difference was found in IL-35 level between the high-risk group and intermediate risk-2 group (P>0.05), but the IL-35 levels in both groups were significantly higher than those in intermediate risk-1 group and the low-risk group (P<0.05), and the level in the intermediate risk-2 group was significantly higher than that in the intermediate risk-1 group and the low-risk group (P<0.05), while there was not different between the intermediate risk-1group and the low risk group (P>0.05). The proportion of iTR35 cells was not significantly different between the high-risk group and the intermediate risk-2 group. The proportions of iTR35 cells in the high-risk group and the intermediate risk-2 group were higher than those in the intermediate risk-1 group and the low-risk group respectively (P<0.05), but there was no differentce in population of iTR35 between the intermediate risk-1 group and the low-risk group (P>0.05). CONCLUSION: The imbalance between IL-35 level and iTR35 cells propertion may play an important role in the development of MDS, which possibly to provides a new theoretical basis for the study of MDS immune targeting therapy.

Octreotide-periplocymarin conjugate prodrug for improving targetability and anti-tumor efficiency: synthesis, in vitro and in vivo evaluation.

Cardiac glycosides could increase intracellular Ca2+ ion by inhibiting the Na+/K+ATPase to induce apoptosis in many tumor cells. However, narrow therapeutic index, poor tumor selectivity and severe cardiovascular toxicity hinder their applications in cancer treatment. To improve the safety profile and tumor targetablility of cardiac glycosides, we designed octreotide conjugated periplocymarin, a cardiac glycoside isolated from Cortex periplocae. The conjugate showed higher cytotoxicity on MCF-7 cells and HepG2 tumor cells (SSTRs overexpression) but much less toxicity in L-02 normal cells. Tissue distribution studies of the conjugate using H22 tumor model in mice showed higher accumulation in tumor and lower distribution in heart and liver than periplocymarin. Furthermore, in vivo anticancer effects of the conjugate on mice bearing H22 cancer xenografts confirmed enhanced anti-tumor efficacy and decreased systemic toxicity. Altogether, octreotide-conjugated periplocymarin demonstrated tumor selectivity and may be useful as a targeting agent to improve the safety profile of cardiac glycosides for cancer therapy.

Ergosterol-loaded poly(lactide-co-glycolide) nanoparticles with enhanced in vitro antitumor activity and oral bioavailability.

AIM: Ergosterol is a plant sterol with anti-tumor and anti-angiogenic activities, but is poorly soluble. In this study, we attempted to enhance its anti-tumor action and oral bioavailability via poly(lactide-co-glycolide) (PLGA) nanoparticle encapsulation. METHODS: Ergosterol-loaded PLGA nanoparticles (NPs/Erg) were prepared using the emulsion/solvent evaporation technique. Their physicochemical properties were characterized, and their cytotoxicity against human cancer cell lines was evaluated with MTT assay. The pharmacokinetics and tissue distribution of NPs/Erg were investigated in rats and mice, respectively. RESULTS: NPs/Erg were spherical in shape with a particle size of 156.9±4.8 nm and a Zeta potential of -19.27±1.13 mV, and had acceptable encapsulation efficiency and loading capacity. NPs/Erg exerted much stronger cytotoxicity against human cancer cells than the free ergosterol, and showed significantly reduced IC50 values (14.69±0.48 µg/mL in glioma U251 cells; 9.43±0.52 µg/mL in breast cancer MCF-7 cells; 4.70±0.41 µg/mL in hepatoma HepG2 cells). After oral administration of a single dose in rats, NPs/Erg displayed a prolonged plasma circulation with a 4.9-fold increase of oral bioavailability compared with the free ergosterol. After mice received NPs/Erg, the ergosterol in NPs/Erg was rapidly distributed in stomach, kidneys, liver, brain, spleen, and virtually non-existent in heart and lungs. The presence of NPs/Erg in brain was particularly improved compared with the free ergosterol. CONCLUSION: The PLGA nanoparticles serve as a promising carrier for the poorly soluble ergosterol and significantly improve its bioavailability, biodistribution and in vitro anti-tumor activities.

Tumor targeted delivery of octreotide-periplogenin conjugate: Synthesis, in vitro and in vivo evaluation.

Periplogenin (PPG), a cardiac glycoside prepared from Cortex periplocae, with similar structure to bufalin, has been found to induce apoptosis in many tumor cells. However, lots of cardiac glycosides possessing strong antitumor activity in vitro have still not passed phase I clinical trials, mostly due to poor tumor selectivity and systemic toxicity. To overcome this drawback, we designed octreotide-periplogenin (OCT-PPG) conjugate by coupling PPG-succinate to the amino-terminal end of octreotide. In comparison with free PPG, the conjugate exhibited significantly stronger cytotoxicity on HepG2 cells (SSTRs overexpression) but much less toxicity in L-02 cells. After intravenous injection of OCT-PPG conjugate into H22 tumor-bearing mice, its total accumulation in tumor was 2.3 fold higher than that of free PPG, but was 0.71- and 0.84-fold lower in heart and liver, respectively, suggesting somatostatin-mediated target delivery of PPG into the tumor tissue and reduced distribution in heart and liver. In vivo studies using H22 tumor model in mice confirmed the remarkable therapeutic effect of this conjugate. These results suggested that OCT-PPG conjugate could provide a new approach for clinical application of cardiac glycosides and as a targeting agent for cancer therapy.

[Expression Analysis and Epigenetics of MicroRNA let-7b in Acute Lymphoblastic Leukemia].

OBJECTIVE: To study the expression and its mechamisms of microRNA let-7b in adult acute lymphoblastic leukemia (ALL), so as to provide the basis for searching a new targeted therapy. METHODS: Firstly, methylation-specific polymerase chain reaction (MSP) was used to analyze the methylation status of CpG islands in microRNA let-7b promoter of bone marrow mononuclear cells in the patients with ALL and patients with non-hematologic malignancies as control, the real-time fluorescence quantitative polymerase chain reaction (qPCR) was used to detect the expression levels of microRNA let-7b in this 2 groups; and then 5-aza-2'-deoxycytidine (5-Aza-dC, DAC) was used to treat ALL cell line MOLT-4; after drug treatment, MSP was used to analyze the methylation status of the CpG islands in microRNA let-7b promoter; the qPCR was used to detect the expression levels of microRNA let-7b, and further explore the regulatory mechanism of microRNA let-7b expression. RESULTS: Hypermethylation of CpG islands in microRNA let-7b promoter in ALL patients was significantly higher than that in patients with non-hematologic malignancies, and the relative expression level of microRNA let-7b was significantly reduced in ALL patients; 5-aza-dC could significantly inhibit the growth of MOLT-4 cells and arrest the cells in G1 phase, thus biosynthesis of RNA and protein was suppressed, and the apoptosis was promoted, meanwhile, 5-Aza-dC could increase the expression of microRNA let-7b. CONCLUSION: In the patients with ALL, the expression of microRNA let-7b is regulated by methylation of CpG islands in the region of genomic promoter. The microRNA let-7b may act as a tumor suppressor, whose low expression is involved in ALL development, indicating the microRNA let-7b may become a new therapeutic target for ALL.

Uniform 2 nm gold nanoparticles supported on iron oxides as active catalysts for CO oxidation reaction: structure-activity relationship.

Uniform Au nanoparticles (∼2 nm) with narrow size-distribution (standard deviation: 0.5-0.6 nm) supported on both hydroxylated (Fe_OH) and dehydrated iron oxide (Fe_O) have been prepared by either deposition-precipitation (DP) or colloidal-deposition (CD) methods. Different structural and textural characterizations were applied to the dried, calcined and used gold-iron oxide samples. Transmission electron microscopy (TEM) and high-resolution TEM (HRTEM) showed high homogeneity in the supported Au nanoparticles. The ex situ and in situ X-ray absorption fine structure (XAFS) characterization monitored the electronic and short-range local structure of active gold species. The synchrotron-based in situ X-ray diffraction (XRD), together with the corresponding temperature-programmed reduction by hydrogen (H2-TPR), indicated a structural evolution of the iron-oxide supports, correlating to their reducibility. An inverse order of catalytic activity between DP (Au/Fe_OH < Au/Fe_O) and CD (Au/Fe_OH > Au/Fe_O) was observed. Effective gold-support interaction results in a high activity for gold nanoparticles, locally generated by the sintering of dispersed Au atoms on the oxide support in the DP synthesis, while a hydroxylated surface favors the reactivity of externally introduced Au nanoparticles on Fe_OH support for the CD approach. This work reveals why differences in the synthetic protocol translate to differences in the catalytic performance of Au/FeOx catalysts with very similar structural characteristics in CO oxidation.

[Lethal effect of cytotoxic lymphocytes against U266 cells induced by DCs modified with GM-CSF gene and pulsed with tumour antigen].

The purpose of this study was to investigate the lethal effect of cytotoxic lymphocytes against U266 cells induced by DCs pulsed with multiple myeloma (MM) U266 lysate and transfected with GM-CSF recombinant adenovirus. The cytotoxic lymphocytes against U266 cells were induced by culturing with DCs, which pulsed with MM U266 antigens and transfected with GM-CSF recombinant adenovirus. The effect of cytotoxic lymphocytes against U266 cells were measured by LDH release detection. Experiments were divided into 3 groups: N-DC group as control in which DCs were normal; U-DC group in which DCs were pulsed by U266 soluble antigen, and G-U-DC group in which DCs were stimulated by U266 soluble antigen and GM-CSF transfected with Ad-CMV. The results showed that there was significant difference on killing rate against U266 cells between 3 groups (F = 10.939, p < 0.05). The killing rate of G-U-DC group was the highest (p < 0.001), and killing rate of U-DC group was higher than that of N-DC group (p < 0.001). It is concluded that the cytotoxic lymphocytes against U266 cells can be induced by DCs pulsed with U266 lysate, and the lethal effect of CTLs can be enhanced when DCs transfected by recombinant adenovirus with exogenous gene GM-CSF.