Changes in Cold and Hot Syndrome and Gastrointestinal Bacterial Community Structure in Mice by Intervention with Food of Different Nature / 中国结合医学杂志

OBJECTIVE@#To reveal the effect of foods with different natures on cold or hot syndrome and gastrointestinal bacterial community structure in mice.@*METHODS@#Forty-five 6-week-old male ICR Kunming mice of clean grade were divided into 5 groups, 9 per group, including the control (CK), hot nature herbs (HM), Hong Qu glutinous rice wine (RW), tea rice wine (TW), and cold nature herbs (CM) groups. Distilled water or corresponding herbs were administered to mice (0.01 mL/g body weight) in the 5 groups by gastric infusion respectively, once daily for 28 d. Appearance, behavior, and serum biochemical indicators, including 5-hydroxytryptamine (5-HT), thyroid stimulating hormone (TSH), noradrenaline (NE), cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), the hot nature index, as well as the gastrointestinal bacterial community structure were analyzed in all groups after treatment.@*RESULTS@#After supplementation for 28 d, CM and TW mice showed different degrees of cold syndrome, and HM and RW mice showed different degrees of hot syndrome. Compared with the HM and RW mice, the TSH, NE, cAMP levels and hot nature indices in the CM and TW mice were significantly decreased and 5-HT and cGMP levels were significantly increased (P<0.05). There was no obvious change in appearance or behavior in CK mice. Results of clustering analysis showed that the gastrointestinal bacterial community structures were highly similar in TW and CM mice as well as in RW and HM mice, and that they were from the same branch, respectively, when the distance was 0.02. The key microbes associated with cold syndrome were Lachnospiraceae uncultured, Lactococcus, etc., and the key microbes associated with hot syndrome were S24-7 norank, Ruminococcaceae uncultured, etc. CONCLUSION: The interventions with different nature foods could change cold or hot syndrome in mice, leading to changes in gastrointestinal bacterial community structure.

Influence of acute pancreatitis in pregnancy on pregnancy outcomes and neonates / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To study the influence of acute pancreatitis in pregnancy (APIP) on pregnancy outcomes and neonates.</p><p><b>METHODS</b>A retrospective analysis was performed for 33 APIP patients and 31 neonates born alive.</p><p><b>RESULTS</b>Of the 33 APIP patients, 26 (79%) developed APIP in the late pregnancy. Fourteen (45%) patients had hyperlipidemic APIP, 13 (42%) had biliary APIP, and 4 (13%) had other types of APIP. According to the severity, 22 (67%) were mild APIP, 5 (15%) were moderate APIP, and 6 were severe APIP. None of the 33 APIP patients died. Among the 20 patients with term delivery, 11 underwent termination of pregnancy; among the 10 patients with preterm delivery, 9 underwent termination of pregnancy; two patients experienced intrauterine fetal death, and one experienced abortion during the second trimester. Among the 31 neonates born alive (two of them were twins), 1 (3%) died, 12 (39%) experienced neonatal hyperbilirubinemia, 8 (26%) had neonatal hypoglycemia, 6 (19%) had neonatal respiratory distress syndrome, 5 (16%) experienced infectious diseases, and 2 (6%) experienced intracranial hemorrhage. The hyperlipidemic APIP group had a higher percentage of patients undergoing termination of pregnancy than the biliary APIP and other types of APIP groups (P<0.05). The incidence rate of preterm infants in the moderate APIP was higher than in the mild and severe APIP groups (P<0.05). The mean birth weights of neonates were the lowest in the moderate APIP group. The incidence rates of neonatal respiratory distress syndrome, intracranial hemorrhage, and infectious disease were the lowest in the mild APIP group (P<0.05).</p><p><b>CONCLUSIONS</b>APIP can lead to adverse pregnancy outcomes and neonatal diseases, which are associated with the severity of pancreatitis.</p>

Screening and validation of long non-coding RNAs in brain tissue of inflammation-induced preterm mice / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To investigate the association between long non-coding RNAs (lncRNAs) and brain injury in inflammation-induced preterm mice, and to provide a reference for the prevention and treatment of brain injury.</p><p><b>METHODS</b>An intraperitoneal injection of lipopolysaccharide in pregnant mice was performed to establish a model of inflammation-induced preterm mice with brain injury (preterm group). The full-term mice delivered by normal pregnant mice were used as controls (full-term group). The lncRNA chip assay was used to screen out the lncRNAs associated with brain injury in preterm mice. Quantitative real-time PCR was used to validate the lncRNAs identified by the above method.</p><p><b>RESULTS</b>The preterm and full-term groups showed significant differences in the expression of 1 978 lncRNAs (P<0.05), consisting of 786 up-regulated lncRNAs and 1 192 down-regulated lncRNAs, and 29 lncRNAs were 1.5 or more times differentially expressed between the two groups. A further analysis was performed for the 10 most differentially expressed lncRNAs, and the results showed that these lncRNAs were involved in the biological processes including transcription, signal transduction, apoptosis, cell cycle, and inflammatory response, as well as G protein-coupled receptor signaling pathway and neuropeptide signaling pathway. Real-time PCR was performed to validate the expression of two lncRNAs in brain tissue in the preterm and full-term groups, and the results were consistent with those of the chip assay.</p><p><b>CONCLUSIONS</b>The expression profiles of lncRNAs in brain tissue change significantly in inflammation-induced preterm mice, and the G protein-coupled receptor signaling pathway may be involved in the pathogenesis of preterm brain injury.</p>

Effects of panaxadiol saponins component as a new Chinese patent medicine on proliferation, differentiation and corresponding gene expression profile of megakaryocytes / 中国结合医学杂志

<p><b>OBJECTIVE</b>To investigate the effects of panaxadiol saponins component (PDS-C) isolated from total saponins of panax ginseng on proliferation, differentiation and corresponding gene expression profile of megakaryocytes.</p><p><b>METHODS</b>Bone marrow culture of colony forming assay of megakaryocytic progenitor cells (CFU-MK) was observed for the promoting proliferation mediated by PDS-C, and differentiation of megakaryocytic blasts caused by PDS-C was analyzed with flow cytometry in CHRF-288 and Meg-01 cells, as well as proliferation, differentiation-related genes expression profile and protein expression levels were detected by human gene expression microarray and western blot.</p><p><b>RESULTS</b>In response to PDS-C 10, 20 and 50 mg/L, CFU-MK from 10 human bone marrow samples was increased by 28.9%±2.7%, 41.0%±3.2% and 40.5%±2.6% over untreated control, respectively (P <0.01, each). Flow cytometry analysis showed that PDS-C treated CHRF-288 cells and Meg-01 cells significantly increased in CD42b, CD41, TSP and CD36 positive ratio, respectively. PDS-C induced 29 genes up-regulated more than two-fold commonly in both cells detected by human expression microarray representing 4000 known genes. The protein expression levels of ZNF91, c-Fos, BTF3a, GATA-1, RGS2, NDRG2 and RUNX1 were increased with western blot in correspond to microarray results.</p><p><b>CONCLUSION</b>PDS-C as an effective component for hematopoiesis, play the role to enhance proliferation and differentiation of megakaryocytes, also up-regulated expression of proliferation, differentiation-related genes and proteins in vitro.</p>

Effects of Danshen Injection () on inhibiting proliferation and inducing apoptosis through down-regulation of mutant JAK2 gene and its protein phosphorylation in human erythroid leukemic cells / 中国结合医学杂志

<p><b>OBJECTIVE</b>To explore the effects of Danshen Injection () on inhibition proliferation, inducing apoptosis and its possible mechanisms on human erythroid leukemic (HEL) cells.</p><p><b>METHODS</b>The commercial Chinese patent medicine of Danshen Injection was extracted and isolated from Chinese herb of Salvia miltiorrhiza bung. The inhibition effects of proliferation were assayed by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) method in HEL cells treated by Danshen Injection at various concentrations for 48 h. The cellular apoptosis was observed in morphology, analyzed by flow cytometry with annexin V and propidium iodide (PI) staining, and examined by DNA degradation ladder on agarose gel electrophoresis. Meanwhile, the expression levels of mutant Janus kinasez (JAK2) gene and phosphorylation-JAK2 (P-JAK2) protein were detected by allele specific-polymerase chain reaction and Western blot.</p><p><b>RESULTS</b>The proliferation of HEL cells was effectively inhibited by Danshen Injection in a dose-dependent manner, with suppression rates from 19.46±2.31% to 50.20±5.21%. Typical apoptosis cells was observed in Danshen Injection treated HEL cells, the rates of annexin V positive cells increased obviously in a dose-dependent manner, as well as the DNA degradation ladder of apoptosis revealed on gel electrophoresis. The expression levels of mutant JAK2 gene and P-JAK2 protein reduced gradually with increasing dosage of Danshen injection.</p><p><b>CONCLUSION</b>Danshen Injection could not only significantly inhibit the proliferation, but also induce apoptosis in HEL cells; down-regulation of the mutant JAK2 gene and P-JAK2 protein expressions are probably one of its molecular mechanisms.</p>

Effects of sodium copper chlorophyllin on mesenchymal stem cell function in aplastic anemia mice / 中国结合医学杂志

<p><b>OBJECTIVE</b>To investigate the effects of sodium copper chlorophyllin (SCC) on the proliferation, differentiation and immunomodulatory function of mesenchymal stem cells (MSCs) from mice with aplastic anemia.</p><p><b>METHODS</b>A mouse model of aplastic anemia was established by exposure of BALB/c mice to sublethal doses of 5.0 Gy Co60 γ radiation, followed by transplantation of 2×10(6) lymph node cells from DBA/2 donor mice within 4 h after radiation. Aplastic anemic BALB/c mice were randomly divided into six groups: the treated groups, which received 25, 50, or 100 mg/kg/day SCC, respectively; a positive control group treated with cyclosporine A (CsA); and an untreated model control group (model group); while, the non-irradiated mice as the normal control group. SCC or CsA were administered by gastrogavage for 20 days, starting on day 4 after irradiation. Peripheral blood cells were counted and colony-forming fibroblasts (CFU-F) in the bone marrow were assayed. The ability of MSCs to form calcium nodes after culture in osteoinductive medium was also observed. The immunosuppressive effect of MSCs on T lymphocytes was analyzed by enzyme-linked immunosorbent assay and flow cytometry, to evaluate the efficacy of SCC in mice with aplastic anemia.</p><p><b>RESULTS</b>Peripheral blood white cell and platelet counts were increased by medium and high SCC doses, compared with the untreated control. CFU-Fs were also increased compared with the untreated control, and the numbers of calcium nodes in MSCs in osteoinductive medium were elevated in response to SCC treatment. The percentage of Forkhead box protein 3 (FOXP3(+)) T cells was increased in T cell-MSC cocultures, and the cytokine transforming growth factor β1 was up-regulated in SCC-treated groups.</p><p><b>CONCLUSION</b>The results of this study suggest that SCC not only promotes the proliferation and differentiation of MSCs, but also improves their immunoregulatory capacity in mice with aplastic anemia.</p>

Panax notoginseng saponins induced up-regulation, phosphorylation and binding activity of MEK, ERK, AKT, PI-3K protein kinases and GATA transcription factors in hematopoietic cells / 中国结合医学杂志

<p><b>OBJECTIVE</b>To investigate the effects of panax notoginseng saponins (PNS) on expression, regulation and phosphorylation of multiple protein kinases in mitogen activated protein kinase (MAPK) intracellular signal pathway and GATA transcription factors in hematopoietic cells, so as to explore its mechanism of proliferation and differentiation activity on hematopoiesis.</p><p><b>METHODS</b>The human granulocytic HL-60, erythrocytic K562, megakaryocytic CHRF-288 and Meg-01 cell lines were treated by PNS, the positive control of K562, CHRF-288 cells treated by recombination human erythropoietin (Epo) and thrombopoietin (Tpo) respectively. The total cell lysate and nuclei protein were extracted after being treated by PNS, subsequently, analyzed by both Western blot and immune-precipitation. Meanwhile, the nuclei extract was performed for electrophoretic mobility shift assay (EMSA) by using (32)P radio labeled double-stranded GATA consensus oligonucleotide.</p><p><b>RESULTS</b>The expression levels of kinase MEK-1, MEK-2, ERK-1, ERK-2, AKT-1, AKT-2 and PI-3K were increased by PNS treatment to different extent in four cell lines, depending on cellular heterogeneity and sensitivity to PNS, also phosphorylation of MEK-1, ERK-1 was differentially promoted by PNS respectively P<0.05, 0.01, 0.001). The expression levels of transcription factors GATA-1 and GATA-2 were increased, moreover, their DNA binding activities were raised dramatically in PNS treated K562, CHRF-288 and Meg-01 cells compared with the controls respectively (P<0.05, 0.01, 0.001). The positive control of K562, CHRF-288 cells treated by Epo or Tpo respectively also displayed up-regulation of protein kinases and GATA transcription factors respectively (P<0.05, 0.01, 0.001).</p><p><b>CONCLUSION</b>The results indicated that intracellular signal pathway initiated by PNS was involved in MAPK pathway and transcription factors of GATA family in hematopoietic cells. PNS displayed the role to promote proliferation and differentiation, by means of increasing expression level and phosphorylation status of multiple protein kinases, also inducing synthesis of GATA transcription factors and upregulation its DNA binding activity.</p>

Effects of Panax notoginseng saponins on proliferation and differentiation in NIH3T3 cells / 中国结合医学杂志

<p><b>OBJECTIVE</b>To investigate the effects of Panax notoginseng saponins (PNS) on the proliferation and differentiation in NIH3T3 cells.</p><p><b>METHODS</b>NIH3T3 cells were treated by various concentrations of PNS 0, 0.05, 0.10, 0.20, and 0.40 g/L. The vitality and proliferation potential of cells were detected by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay, the alkaline phosphatase (ALP) activity was measured by p-nitrophenyl phosphate (pNPP) assay, and the mineralization formation ability was tested for the cellular differentiation toward osteoblast, as well as the expression level of phosphorylated extracellular signal-regulated kinase1/2(P-ERK1/2), extracellular signal-regulated kinase1/2 (ERK1/2) protein kinase was analyzed by Western blot with total cell lysate of NIH3T3 cells treated by PNS.</p><p><b>RESULTS</b>Both MTT and pNPP assay showed that optical density (OD) values were increased in response to PNS treatment at a dose-dependent pattern. The mineralization formation ability was enhanced in PNS-treated NIH3T3 cells compared with untreated cells. Meanwhile, the expression level of P-ERK1/2 protein kinase was up-regulated in PNS-treated NIH3T3 cells, while, the expression level of ERK1/2 protein kinase revealed no obvious difference with or without PNS treated cells.</p><p><b>CONCLUSION</b>PNS could pay a role to promote the proliferation and differentiation in NIH3T3 cells by means of up-regulation of P-ERK1/2 protein kinase.</p>

Experimental study of hematopoietic cell gene expression profile induced by panax notoginosides in vitro / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To explore the relationship of up-regulated genes with cell proliferation and differentiation by analyzing the hematopoietic cells gene expression profile induced by panax notoginosides (PNS) using cDNA microarray.</p><p><b>METHODS</b>The cDNA membrane microarray with 480 target genes related to proliferation and differentiation of hematopoietic cells was prepared, and mRNA was extracted and purified from 3 lineages of human hematopoietic cell lines, including megakaryocytic CHRF-288, granulocytic HL-60 and erythrocytic K562 cells, respectively after they were treated with PNS. The hybridization with target genes on microarray membrane was performed using [alpha-33 ]dATP labeled cDNA from reversed mRNA.</p><p><b>RESULTS</b>After treated by PNS, the genes up-regulated for more than 3 folds could be classified to 11 sorts according to their function, including the methyl-transferase, acetyl-transferase, differentiation initiated factor, anti-apoptosis, transcription regulation protein, cell cycle related protein, protein and kinase of signal pathway, receptors, DNA or RNA polymerase, protein phosphatase, transporter or trafficking protein and rat sarcoma (RAS) homology gene family. In three cell lines of CHRF-288, HL-60 and K562 treated by PNS, 78 (16.3%), 89 (18.5%) and 59 (12.3%) pieces of genes respectively were up-regulated for more than 3 folds.</p><p><b>CONCLUSION</b>All the up-regulated genes induced by PNS in microarray analysis were related to hematopoietic cell proliferation and differentiation, the outcome is in accord with the results reported previously by the authors from the studies of mice model with hematopathy, hematopoietic stem/progenitor cells, gene transcription regulation and protein kinase of signal pathway, etc. It provides a powerful evidence for the PNS activity and its mechanism by gene expression profile.</p>

Effects of Ginseng panaxadiol saponin on proliferation and differentiation of human bone marrow CD34+ cells / 中国实验血液学杂志

The study was purposed to investigate the effects of the panaxadiol saponin (PDS) from Ginseng on proliferation and differentiation of human CD34(+) cells from human bone marrow. Highly purified CD34(+) cells were isolated from human bone marrow by using the Dynal CD34 Cell Selection System (Dynal, Norway). The cells were exposed to PDS at various concentrations in both agar semi-solid culture of CFU-Mix and suspension culture of myeloid and erythroid cells in order to observe the effects of PDS on proliferation of CD34(+) cells. The cells were marked with 4 kinds of monoclonal antibody in related with their differentiation toward to myeloid and erythroid lineages, then examined by flow cytometry (FACS) after being incubated with PDS for 14 days. The results showed that the number of CD34(+) cells was 1.0 +/- 0.15% out of marrow nuclear cells after being purified by Dynal beads system. The enrichment of CD34(+) cells reached to 86.8 +/- 2.8%. The best efficiency in promoting proliferation of CD34(+) cells in vitro was obtained when the concentration of PDS was 25 mg/L, the formation of CFU-Mix colonies significantly increased by 56.3 +/- 3.5% over those of no-PDS control (p < 0.01). The results from suspension culture revealed that myeloid cells elevated in a dose-dependent manner with a peak increasing rate of 35.6 +/- 3.2%, and erythroid cells significantly increased by 22.3 +/- 2.1% over those of no-PDS control (all p < 0.01). After incubation with PDS for 14 days, number of CD33(+) cells increased in a dose-dependent manner with a peak increasing rate at 50 mg/L. CD71(+) cells reaching the peak were at 25 mg/L, while G-A(+) cells were increased by 7.2 +/- 1.3% (p < 0.01) at 10 mg/L, but the number of CD15(+) cells was not found to be changed before and after treating with PDS. It is concluded that PDS not only enhance the proliferation of CD34(+) cells, but also induce differentiation of CD34(+) cells toward to myeloid and erythroid lineages. PDS may play the roles as like hematopoietic growth factor, or provide synergistic effects on growth factor in the regulation of hematopoiesis.

Up-regulation of transcription factors GATA-1 and GATA-2 induced by Panax notoginosides in hematopoietic cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To observe the role of Panax notoginosides (PNS) in up-regulation of GATA family transcription factors, and explore intracellular signal pathway of PNS in the proliferation of hematopoietic cells.</p><p><b>METHODS</b>Human bone marrow cells were incubated with different concentrations of PNS for colony-forming assay. Human cell lines HL-60, K562, CHRF-288 and Meg-01 were incubated with PNS (10 mg/L) for 14 days. The cell nuclear proteins were extracted and analyzed by Western blot with antibodies against GATA-1, GATA-2. Electrophoretic mobility shift assay (EMSA) and antibody gel supershift assay was performed using (32)P labeled GATA consensus oligonucleotide which contains binding site for GATA transcription factors.</p><p><b>RESULTS</b>PNS could promote the proliferation of CFU-GM and CFU-E and induce the expression of GATA-1, GATA-2. The nuclear proteins of both GATA-1 and GATA-2 in K562, CHRF-288 and Meg-01 cells treated by PNS were increased by (1.5 - 2.8) and (2.0 - 3.1)-fold over untreated cells respectively. GATA binding activity initiated by PNS was apparently elevated to form higher density band of GATA-DNA complex. While there was no detectable change in HL-60 cells before and after PNS treatment. The predominant GATA binding complex was mainly attributable to both GATA-1 and GATA-2 proteins being in phosphorylated status.</p><p><b>CONCLUSION</b>PNS can induce the synthesis of transcription factors GATA-1 and GATA-2 and enhance their DNA binding activity, which could play a role in the up-regulation of the expression genes related to proliferation and differentiation in hematopoietic cells.</p>