OBJECTIVE: To study the effect of taspine hydrochloride (TA/HCl) on skin wound healing in rats and its mechanism. METHODS: Bilateral round wounds were made on the backs of SD rats. The effect of TA/HCl on the skin wound was evaluated through determining closure time and contracting ability of the skin wound, observing histopathological characteristics and measuring contents of hydroxyproline (Hyp) and protein in the wound tissue. RESULTS: The closure time of the skin wounds was significantly shorter in the TA/HCl-treated groups than that in the model group. The percentages of wound contraction were higher in the TA/HCl-treated groups than that in the dimethyl sulfoxide (DMSO) control group of the same group (P<0.05 or P<0.01) on the 3rd to 14th days after wounding. The content of the protein in the wound tissue in the TA/HCl-treated group (2 mg/ml) was higher than that in the model group (P<0.05) on the 3rd to 7th days after wounding, and it arrived at the peak on the 7th day and gradually decreased to the normal level in skin tissue on the 14th to 21st days after wounding. The contents of Hyp in the wound tissues in the TA/HCl-treated groups were higher than that in the model group (P<0.05 or P<0.015) on the 3rd to 21st days after wounding, and they arrived at the peak on the 14th day and at the normal level in skin tissue on the 21st day. Histopathological test results showed that TA/HCl could promote the formation of newly born capillaries in the early period of the wound healing. CONCLUSION: TA/HCl has the ability of promoting skin wound healing in rats, and it can also accelerate the growth of newly born capillaries and raise the production of protein and collagen in wound tissue.

Objective To investigate the biological characteristics and epidemiological significance of Yersinia pestis strains in Qilian County,Qinghai Province,in order to provide a scientific basis for plague prevention and control.Method Totally 67 strains were separated from kinds of host in Qilian County,Qinghai Province from 1958 to 2011,to do biochemical test,toxicity test,virulence factors evaluation,plasmid analysis and different region (DFR) genotyping.Results According to biochemical typing,48 of the 50 strains tested were Qing-Tibet Plateau ecotype,15 were Qilian Mountain ecotype,and the remaining 4 were different ecotypes from the plague foci in Qinghai plateau.The strains had 8 genomovars,and were given priority to genomovar8 (42 strains),secondly,genomovar44 (15 strains),genomovar5 (4 strains),genomovar7 (2 strains),genomovar19 (1 strain),genomovar30 (1 strain),genomovar32 (1 strain),and genomovar34 (1 strain).A proportion of 95.52％ (64/67) of the strains contained 3 kinds of plasmid-6 × 106,45 × 106,and 52 × 106;85.07％ (57/67) contained all the four virulence factors,and 96.00％ (48/50) were velogenic strains.Conclusion The strains separated in Qilian County,Qinghai Province have the characteristics of Qinghai-Tibet Plateau plague's pathogen and have strong toxicity,so we should enhance the plague monitoring and give more publicity to plague prevention to prevent animal plague spreading to human.

OBJECTIVETo explore the interaction between P311 and transforming growth factor beta 1 (TGF-β1) in murine fibroblasts and its effect on the function of fibroblasts.

METHODSSkin fibroblasts obtained from five neonatal P311 wild-type C57BL/6 mice and P311 gene knock-out C57BL/6 mice were cultured. The second passage of fibroblasts were used in the following experiments. All experiments were repeated for 3 times. (1) The fibroblasts of P311 wild-type mice were divided into blank control group and P311 over-expression group according to the random number table (the same grouping method below), with 36 wells in each group. Fibroblasts in blank control group were transfected with 10 μL control vector, and fibroblasts in P311 over-expression group were transfected with equal efficiency P311 expression adenovirus vector. After being cultured for 48 hours, the mRNA expression level of P311, and the mRNA and protein expression levels of TGF-β1, α-smooth muscle actin (α-SMA), and collagen type I of fibroblasts in both groups were determined with real-time fluorescent quantitative RT-PCR and Western blotting (the same detection methods below), respectively. (2) After cultured reaching the cell density of 80%-90%, the mRNA and protein expression levels of TGF-β1, α-SMA, and collagen type I of the fibroblasts of P311 wild-type mice and P311 gene knock-out mice, with 4 flasks in each type of fibroblasts, were determined. (3) The fibroblasts of P311 wild-type mice were divided into blank control group and 5, 10, 15, 20, and 25 ng/mL TGF-β1 groups after being starved treatment with DMEM medium containing 1% FBS for 3 hours, with 2 flasks in each group. Fibroblasts in blank control group were routinely cultured, while fibroblasts in the latter five groups were treated with 5, 10, 15, 20, and 25 ng/mL TGF-β1, respectively. After being cultured for 48 hours, the mRNA expression levels of P311 in fibroblasts of the six groups were determined. Another fibroblasts of P311 wild-type mice were divided into blank control group and 10 ng/mL TGF-β1 group, with 6 wells in each group, and the protein expression levels of P311 in both groups were determined by immunofluorescence staining. (4) The fibroblasts of P311 wild-type mice were divided into blank control group and 10 ng/mL TGF-β1 group after being starved treatment as above, with 2 flasks in each group, and fibroblasts in blank control group were routinely cultured, while fibroblasts in the latter group were treated with 10 ng/mL TGF-β1. After being cultured for 48 hours, the mRNA and protein expression levels of α-SMA and collagen type Ⅰ were determined. The fibroblasts of P311 gene knock-out mice were grouped and treated as above, and the mRNA and protein expression levels of α-SMA and collagen type I were determined. Data were processed with one-way analysis of variance and t test.

RESULTS(1) The mRNA expression level of P311 of fibroblasts in P311 over-expression group was increased nearly 300 000-fold compared with that in blank control group (t=9.942, P<0.001). The mRNA expression levels of TGF-β1, α-SMA, and collagen type I of fibroblasts in P311 over-expression group, and the protein expression levels of pro-TGF-β1, activated TGF-β1, α-SMA, and collagen type I of fibroblasts in P311 over-expression group were significantly higher than those in blank control group (with t values from 8.192 to 49.090, P values below 0.01). (2) The mRNA expression levels of TGF-β1, α-SMA, and collagen type I in fibroblasts of P311gene knock-out mice were significantly lower than those in fibroblasts of P311 wild-type mice (with t values from 8.157 to 22.270, P values below 0.01). The protein expression levels of pro-TGF-β1, α-SMA, and collagen type I in fibroblasts of P311 gene knock-out mice were significantly lower than those in fibroblasts of P311 wild-type mice (with t values from 2.995 to 12.600, P<0.05 or P<0.01), and the protein expression levels of active TGF-β1 were similar in two types of fibroblasts (t=1.070, P>0.05). (3) The mRNA expression levels of P311 of fibroblasts in blank control group and 5, 10, 15, 20 and 25 ng/mL TGF-β1 groups were 1.28 ± 0.44, 3.61 ± 0.91, 6.64 ± 0.92, 6.58 ± 1.04, 1.79 ± 0.31, 0.16 ± 0.06, respectively. Compared to the mRNA expression level of P311 of fibroblasts in the blank control group, the mRNA expression levels of P311 of fibroblasts in 5 and 20 ng/mL TGF-β1 groups were similar (with t values respectively 2.302 and 0.955, P values above 0.05), while they were significantly higher in 10 and 15 ng/mL TGF-β1 groups (with t values respectively 5.630 and 4.710, P values below 0.001), and they were significantly lower in 25 ng/mL TGF-β1 group (t=2.509, P<0.01). The protein expression level of P311 of fibroblasts in 10 ng/mL group was higher than that in blank control group. (4) The mRNA and protein expression levels of α-SMA and collagen type I of fibroblasts of P311 wild-type mice in 10 ng/mL TGF-β1 group were significantly higher than those in blank control group (with t values from 3.523 to 14.290, P<0.05 or P<0.01). The mRNA and protein expression levels of α-SMA and collagen type I of fibroblasts of P311 gene knock-out mice in 10 ng/mL TGF-β1 group were significantly higher than those in blank control group (with t values from 4.895 to 14.870, P<0.05 or P<0.01).

CONCLUSIONSThe interaction between P311 and TGF-β1 in murine fibroblasts exists and it may enhance the differentiation of fibroblasts in combination.

Actins ; metabolism ; Animals ; Cell Differentiation ; Cells, Cultured ; Collagen Type I ; metabolism ; Fibroblasts ; cytology ; metabolism ; Mice ; Mice, Inbred C57BL ; Mice, Knockout ; Nerve Tissue Proteins ; genetics ; metabolism ; RNA, Messenger ; Transfection ; Transforming Growth Factor beta1 ; metabolism

Objective To evaluate the efficacy and safety of intensity modulated radiation therapy (IMRT) for whole-brain radiotherapy with simultaneous integrated boost for brain metastases.Methods From January 2015 to January 2017,120 patients with brain metastases undergoing hospital radiotherapy at the First People's Hospital and the Second People's Hospital of Yulin City of Guangxi Zhuang Autonomous Region were randomly divided into three dimensional conformal radiation therapy (3DCRT) group (n =60) and IMRT group (n =60) by envelope method.The curative effects of the two groups and the changes of patient's mental states before and after radiotherapy were compared.Results The median survival times of patients in 3DCRT group was 11.5 months,and the 6 months and 1 year survival times rates were 83.3％ and 35.0％ respectively.The median survival times of patients in IMRT group was 12.9 months,and the 6 months and 1 year survival rates were 86.7％ and 38.3％ respectively.There was no significant difference in prognosis between the two groups (x2 =0.143,P =0.705).There were 8 cases with complete remission,38 cases with partial remission,11 cases with stable disease,3 cases with progressive disease in 3DCRT group,and there were 9,40,9,2 cases in IMRT group respectively.There was no statistically significant difference in short-term efficacy between the two groups (Z =-0.641,P =0.520).Univariate analysis results showed that recursive partitioning analysis (RPA) classification (x2=53.484,P ＜0.001),extracranial lesions control (x2=13.392,P ＜ 0.001),whether the midline offset (x2 =4.427,P =0.035) were prognostic factors.Multivariate analysis results showed that RPA classification (HR =2.631,95 ％ CI:1.884-3.673,P ＜ 0.001),extracranial lesions control (HR =1.697,95％ CI:1.034-2.786,P =0.037),whether the midline offset (HR =1.787,95％ CI:1.118-2.856,P =0.015) were the independent prognostic factors for the overall survival.According to the results of activities of mini-mental state examination,the scores of 3DCRT group before and after radiotherapy were 27.12 ± 2.42 and 26.08 ± 2.42 respectively,with a significant difference (t =2.723,P =0.009);and the scores of IMRT group before and after radiotherapy were 26.57 ± 3.09 and 27.20 ± 2.46 respectively,with no significant difference (t =-1.610,P =0.1 13).Conclusion IMRT for whole-brain radiotherapy with simultaneous integrated boost is an effective treatment for multiple brain metastases,which can relieve symptoms,prolong survival time and improve quality of life.

Objective To establishment a method for detection of multiple drug resistance gene of Yersina pestis using polymerase chain reaction(PCR), to provide a guidance for treatment of plague. Methods According to National Center for Biotechnology Information (NCBI) released sequences of aminoglycoside resistant genes of streptomycin resistant,strB,strA,beta lactam antibiotics resistant genes tem,shv,and ctx-m,sulfamilamide resistant genes sul1, sul2, and sul3, a pair of primers of each gene was designed. DNAs of 282 strains isolated from plague natural foci in Qinghai Province were amplified by PCR using every pair of primers. The products were separated using gel electrophoresis, and the results were visualized through a gel imaging system. The susceptibility of 282 Yersina pestis to streptomycin, sulfamethoxazole and ceftriaxone was tested by drug sensitivity test. Results The PCR amplification results of all samples were negative,and strains with streptomycin,sulfamilamide and beta lactam antimicrobial drug resistance genes were not found. Drug sensitivity test showed that 282 strains were highly sensitive to streptomycin,sulfamethoxazole and ceftriaxone sodium.The diameter of bacteriostasis ring>19,17,21 mm, respectively. Conclusions It is a feasible method to use PCR technology to detect the multiple drug resistance genes of Yersinia pestis. Using this method to systematically monitor the resistance gene of Yersinia pestis is an efficient, economical and practical experimental method, which can provide guidance for the treatment of plague disease.

Objective: To explore the effects of hypoxia inducible factor-1α (HIF-1α) on P311 and its influence on the migration of murine epidermal stem cells (ESCs) under hypoxia in vitro. Methods: Two kinds of murine ESCs were isolated and obtained from 15 neonatal wild-type C57BL/6J mice and 5 congeneric source P311 gene knock-out mice, respectively. The first passage of cells were used in the following experiments after morphologic observation and detection of expression of cell surface markers CD71 and CD49f with flow cytometer. (1) After cell scratch assay, according to the random number table (the same dividing method below), ESCs of P311 gene knock-out mice were divided into normoxia group (cells were cultured with complete medium in normoxic carbon dioxide incubator, and the subsequent normoxic treatments were the same) and hypoxia group (cells were cultured in hypoxic carbon dioxide incubator containing 1% oxygen, and the subsequent hypoxic treatments were the same), with 12 inserts in each group. ESCs of wild-type mice were divided into normoxia group, pure hypoxia group, dimethyl sulfoxide (DMSO) control group (2 μL DMSO solvent was added for 1 h of normoxia treatment before hypoxia treatment), HIF-1α inhibitor group (cells were treated with 11 μmol/L HIF-1 inhibitor of 2 μL under normoxia condition for 1 h before hypoxia treatment), HIF-1α stabilizer group (the cells were treated with 2 μmol/L FG-4592 of 2 μL under normoxia condition for 1 h before hypoxia treatment), with 12 inserts in each group. Three inserts of each time point in each group were adopted respectively to measure the residual width of scratch under inverted phase contrast microscope at post scratch hour (PSH) 0 (immediately), 12, 24, and 48. (2) After hypoxia treatment, the protein level of HIF-1α in ESCs of wild-type mice was detected by Western blotting at post hypoxia hour (PHH) 0, 12, 24, and 48. (3) ESCs of wild-type mice were divided into pure hypoxia group, DMSO control group, HIF-1α inhibitor group, and HIF-1α stabilizer group as that of experiment (1) with the same treatment. The mRNA expression of P311 and expression of P311 in ESCs were determined by real-time fluorescent quantitative reverse transcription polymerase chain reaction and immunocytochemical staining, respectively, at PHH 0 (immediately), 12, 24, and 48 (with sample numbers of 12). (4) The second passage of human embryonic kidney 293 (HEK-293) cells were divided into empty vector hypoxia group (cells were cultured under hypoxia condition after being transfected with empty vector plasmid), P311 normoxia group (cells were cultured under normoxia condition after being transfected with P311 reporter gene plasmid), P311 hypoxia group (cells were cultured under hypoxia condition after being transfected with P311 reporter gene plasmid), P311 hypoxia+ HIF-1α inhibitor group (cells which were incubated with HIF-1α inhibitor were cultured under hypoxia condition after being transfected with P311 reporter gene plasmid). The luciferase activity was detected at post culture hour (PCH) 0 and 12, respectively, and then the P311 transcriptional regulatory binding site of HIF-1α and the promoter sequence of P311 were predicted and searched by bioinformatics methods. Data were processed with factorial design variance analysis, one-way analysis of variance, LSD test and Bonferroni correction. Results: (1) The results of ESCs. The cells showed cobblestone-like pattern and different clonal morphology due to the different cell proliferation potential. The proportion of CD71^-CD49f⁺ cells accounted for about 85%. The identification results indicated that the cells showed strong stem cell properties and high purity. Compared with those in cells of normoxia group of P311 gene knock-out mice, the residual widths of scratch of cells in pure hypoxia group were smaller at PSH 12 and 24 (with P values below 0.05), and those in hypoxia group, normoxia group of wild-type mice, DMSO control group, HIF-1α inhibitor group, and HIF-1α stabilizer group were smaller at PSH 12 (with P values below 0.05). Compared with those in cells of normoxia group of wild-type mice, the residual widths of scratch of cells in hypoxia group, pure hypoxia group, and DMSO control group were smaller at PSH 12 and 24 (with P values below 0.05), and the residual width of scratch of cells in HIF-1α stabilizer group was smaller at PSH 12 (P<0.05). Compared with those of cells in pure hypoxia group, the residual widths of scratch of cells in hypoxia group were wider at PSH 12 and 24 (with P values below 0.05), and the residual width of scratch of cells in HIF-1α inhibitor group was wider at PSH 12 (P<0.05), and those of cells in HIF-1α stabilizer group were smaller at PSH 12 and 24 (with P values below 0.05). There was no obvious difference in the width of scratch in cells among the 7 groups (F=19.02, P>0.05). The protein levels of HIF-1α in ESCs of wild-type mice at PHH 0, 12, 24, and 48 were respectively 1.02±0.05, 2.56±0.09, 1.60±0.17, and 1.17±0.03. Compared with that at PHH 0, the protein level of HIF-1α at PHH 12 was significantly enhanced (P<0.01). It began to decline at PHH 24 but was still higher than that at PHH 0 (P<0.05), and the protein level of HIF-1α at PHH 48 was close to the normoxia level (P>0.05). Compared with those of cells in pure hypoxia group, the mRNA expressions of P311 of cells in HIF-1α inhibitor group were significantly decreased at each time point (with P values below 0.05), and those in HIF-1α stabilizer group were significantly increased at PHH 12 and 24 (with P values below 0.05). Compared with those of cells in HIF-1α inhibitor group, the mRNA expressions of P311 of cells in DMSO control group and HIF-1α stabilizer group were significantly increased at PHH 0, 12, and 24 (with P values below 0.05). Compared with those of cells in pure hypoxia group, the expressions of P311 of cells in HIF-1α inhibitor group were significantly decreased at each time point (with P values below 0.05), while those in HIF-1α stabilizer group were significantly increased at PHH 12 and 24 (with P values below 0.05). Compared with those of cells in HIF-1α inhibitor group, the expressions of P311 of cells in DMSO control group and HIF-1α stabilizer group were significantly increased at PHH 12 and 24 (with P values below 0.05). (2) The results of HEK-293 cells. At PCH 0, there was no significant difference in the luciferase activity among cells of empty vector hypoxia group, P311 normoxia group, P311 hypoxia group, and P311 hypoxia+ HIF-1α inhibitor group (F=13.33, P>0.05). At PCH 12, the luciferase activity of cells in P311 hypoxia group was higher than that in empty vector hypoxia group (P<0.01). The luciferase activity of cells in hypoxia group was higher than that in P311 normoxia group (P<0.05), while that of cells in P311 hypoxia+ HIF-1α inhibitor group was lower than that in P311 hypoxia group (P<0.01). Conclusions HIF-1α may increase the migration of murine ESCs through inducing the expression of P311 at the early stage of hypoxia.

This study aims to explore the diagnostic value of specific immunoglobulin E (sIgE) and specific immunoglobulin G (sIgG) of Aspergillus fumigatus in the diagnosis of allergic broncho-pulmonary aspergillosis (ABPA) and severe asthma with fungal sensitization (SAFS). A total of 17 ABPA patients and 14 SAFS patients were enrolled. The levels of sIgG [2 294.00 (1 527.00, 14 170.00) U/ml vs. 972.60 (650.90, 1 792.00) U/ml] and sIgE [8.77 (1.64, 16.85) kU/L vs. 1.04 (0.70, 2.05) kU/L] in ABPA patients were significantly higher than those in SAFS patients (P<0.05). Aspergillus fumigatus sIgG was strongly correlated with Aspergillus fumigatus sIgE (r_s=0.797, P<0.001) in ABPA patients. When combined with Aspergillus fumigatus sIgG (>1 000.00 U/mL) and Aspergillus fumigatus sIgE (>1.00 kU/L), the sensitivity was 82.3% and specificity 78.6% for the differential diagnosis of ABPA and SAFS. It demonstrates the diagnostic value of Aspergillus fumigatus sIgG and sIgE.

Liposomes mimic natural cell membranes and have long been investigated as drug carriers due to excellent entrapment capacity, biocompatibility and safety. Despite the success of parenteral liposomes, oral delivery of liposomes is impeded by various barriers such as instability in the gastrointestinal tract, difficulties in crossing biomembranes, and mass production problems. By modulating the compositions of the lipid bilayers and adding polymers or ligands, both the stability and permeability of liposomes can be greatly improved for oral drug delivery. This review provides an overview of the challenges and current approaches toward the oral delivery of liposomes.

Biomimetic nanocarriers are emerging as efficient vehicles to facilitate dietary absorption of biomacromolecules. In this study, two vitamins, thiamine and niacin, are employed to decorate liposomes loaded with insulin, thus facilitating oral absorption vitamin ligand-receptor interactions. Both vitamins are conjugated with stearamine, which works to anchor the ligands to the surface of liposomes. Liposomes prepared under optimum conditions have a mean particle size of 125-150 nm and an insulin entrapment efficiency of approximately 30%-36%. Encapsulation into liposomes helps to stabilize insulin due to improved resistance against enzymatic disruption, with 60% and 80% of the insulin left after 4 h when incubated in simulated gastric and intestinal fluids, respectively, whereas non-encapsulated insulin is broken down completely at 0.5 h. Preservation of insulin bioactivity against preparative stresses is validated by intra-peritoneal injection of insulin after release from various liposomes using the surfactant Triton X-100. In a diabetic rat model chemically induced by streptozotocin, both thiamine- and niacin-decorated liposomes showed a comparable and sustained mild hypoglycemic effect. The superiority of decorated liposomes over conventional liposomes highlights the contribution of vitamin ligands. It is concluded that decoration of liposomes with thiamine or niacin facilitates interactions with gastrointestinal vitamin receptors and thereby facilitates oral absorption of insulin-loaded liposomes.

The long-circulating effect is revisited by simultaneous monitoring of the drug payloads and nanocarriers following intravenous administration of doxorubicin (DOX)-loaded methoxy polyethylene glycol-polycaprolactone (mPEG-PCL) nanoparticles. Comparison of the kinetic profiles of both DOX and nanocarriers verifies the long-circulating effect, though of limited degree, as a result of pegylation. The nanocarrier profiles display fast clearance from the blood despite dense PEG decoration; DOX is cleared faster than the nanocarriers. The nanocarriers circulate longer than DOX in the blood, suggesting possible leakage of DOX from the nanocarriers. Hepatic accumulation is the highest among all organs and tissues investigated, which however is reversely proportionate to blood circulation time. Pegylation and reduction in particle size prove to extend circulation of drug nanocarriers in the blood with simultaneous decrease in uptake by various organs of the mononuclear phagocytic system. It is concluded that the long-circulating effect of mPEG-PCL nanoparticles is reconfirmed by monitoring of either DOX or the nanocarriers, but the faster clearance of DOX suggests possible leakage of a fraction of the payloads. The findings of this study are of potential translational significance in design of nanocarriers towards optimization of both therapeutic and toxic effects.