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OBJECTIVE To investigate the improvement effects of Runchang granules on the constipation in mice and its potential mechanism. METHODS The mice were randomly divided into normal control group， model group， Runchang granules low-dose and high-dose groups （5， 10 g/kg）， mosapride group （0.003 g/kg， positive control）， with 6 mice in each group. The latter 4 groups were given loperamide intragastrically （0.004 g/kg）， twice a day， for 3 consecutive days. Normal control group and model group were given purified water intragastrically， and administration groups were given relevant medicine intragastrically for 7 consecutive days. After the last medication， fecal moisture content and intestinal motility of mice were determined， while the structures of colon and ileum， and the secretion of colonic mucus were observed. Protein expressions of tyrosine kinase receptor （c-kit）， mucin 2 （MUC2） and stem cell factor （SCF） were determined in colon； meanwhile， the mRNA expression levels of inflammatory factors [tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6）， IL-1β， inducible nitric oxide synthase （iNOS）] as well as factors related to promoting intestinal motility [neuronal nitric oxide synthase （nNOS）， smooth muscle myosin light chain kinase （smMLCK）， 5-hydroxytryptamine 4 receptor （5-HT4R）， MUC2， SCF， c-kit] were determined. RESULTS Compared with model group， the fecal water content， intestinal propulsion rate， protein expression of c-kit in colon， relative expressions of MUC2 and SCF protein， and mRNA expressions of factors related to promoting intestinal motility （except for nNOS and SCF in Runchang granules low-dose group） were all increased significantly in Runchang granules low-dose and high-dose groups， and mosapride group （P＜0.05 or P＜0.01）. mRNA expression levels of inflammatory factors were decreased significantly（P＜0.05 or P＜0.01）. Both colon and ileum injuries improved， and the secretion of colon mucus was increased significantly in Runchang granules high-dose group （P＜0.01）. CONCLUSIONS Runchang granules have laxative effect and can improve constipation in mice， and its mechanism may be related to the promotion of the secretion of colon mucus and MUC2 expression， and the activation of SCF/c-kit signaling pathway.
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Objective@#To study the effect of stem cell factor (SCF) on the angiogenic ability of cocultured dental pulp stem cells (DPSCs) and human umbilical vein endothelial cells (HUVECs).@*Methods @#This study has been reviewed and approved by the Ethics Committee. The experiment was split into the HUVECs, SCF+HUVECs, DPSCs+HUVECs, and SCF+DPSCs+HUVECs groups. A mixture of SCF and culture medium was used to prepare a mixed culture medium with an SCF concentration of 100 ng/mL. In vitro coculture of DPSCs and HUVECs was performed at a 1∶5 ratio. CCK-8 proliferation assay was used to observe the proliferative capacity of cells in each group on days 1, 3, 5, and 7. Wound healing and Transwell migration assays were used to detect the effect of SCF on cell migration under either direct or indirect coculture conditions, respectively. In vitro angiogenesis experiments were performed to detect the angiogenic capacity of the cells in each group. The vascular endothelial growth factor A (VEGFA) concentration in the cell culture supernatant was detected using ELISAs, and the protein expression levels of CD31, CD34, and VEGFA were detected using Western blot analysis. @*Results @# Wound healing and Transwell migration experiments showed that SCF significantly promoted the migration of cocultured DPSCs and HUVECs (P<0.05). The in vitro angiogenesis experiment showed that the number of branches and the total length of branches of tubular structures in the SCF+DPSCs+HUVECs group were significantly greater than those of the other groups (P<0.05), and the expression levels of the vascular-related proteins CD31, CD34, and VEGFA in this group were greater (P<0.01). @*Conclusion @# SCF can enhance the migration and in vitro angiogenesis of cocultured DPSCs and HUVECs.
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OBJECTIVE@#To observe the effects of moxibustion on the stem cell factor (SCF)/tyrosine kinase receptor (c-kit) signaling pathway and immune function in rats with diarrhea irritable bowel syndrome (IBS-D), and to explore the mechanism of moxibustion for IBS-D.@*METHODS@#Among 52 young rats born from 6 healthy pregnant SPF rats, 12 rats were randomly selected into the normal group, and the remaining 40 rats were treated with the three-factor combination method of maternal separation, acetic acid enema and chronic restraint stress to establish the IBS-D rat model. Thirty-six rats with successful IBS-D model were randomly divided into a model group, a moxibustion group, and a medication group, 12 rats in each group. The rats in the moxibustion group were treated with suspension moxibustion at "Tianshu" (ST 25) and "Shangjuxu" (ST 37); the rats in the medication group were treated with intragastric administration of rifaximin suspension (150 mg/kg). All the treatments were given once a day for 7 consecutive days. The body mass, loose stool rate (LSR), the minimum volume threshold when abdominal withdrawal reflex (AWR) scored 3 were measured before acetic acid enema (35 days old), after modeling (45 days old), and after intervention (53 days old). After intervention (53 days old), HE staining was used to observe the morphology of colon tissue, and spleen and thymus coefficients were measured; ELISA method was used to detect serum inflammatory factors (tumor necrosis factor a [TNF-a], interleukin [IL]-10, IL-8), T-lymphocyte subsets (CD+4, CD+8, CD+45), value of CD+4/CD+8 and immune globulin (IgA, IgG, IgM); real-time PCR method and Western blot method was used to detect the expression of SCF, c-kit mRNA and protein in colon tissue; immunofluorescence staining method were used to detect positive expression of SCF and c-kit.@*RESULTS@#After intervention, compared with the normal group, in the model group, the body mass and the minimum volume threshold when AWR scored 3 were decreased (P<0.01), LSR, spleen and thymus coefficients, serum levels of TNF-α, IL-8, CD+4, CD+45, CD+4/CD+8, IgA, IgG, IgM were increased (P<0.01), serum IL-10 level and protein and mRNA expression of SCF and c-kit in colon tissue were decreased (P<0.01), and the positive expression of SCF and c-kit was decreased (P<0.01). Compared with the model group, in the moxibustion group and the medication group, the body mass and the minimum volume threshold when AWR scored 3 were increased (P<0.01, P<0.05), LSR, spleen and thymus coefficients, serum levels of TNF-α, IL-8, CD+4, CD+8, CD+45, CD+4/CD+8, IgA, IgG, IgM were decreased (P<0.01, P<0.05), serum IL-10 level and protein and mRNA expression of SCF and c-kit in colon tissue were increased (P<0.01), and the positive expression of SCF and c-kit was increased (P<0.01). Compared with the medication group, in the moxibustion group, the level of serum CD+4 was decreased (P<0.05), the value of CD+4/CD+8 was increased (P<0.01), and there was no significant difference in other indexes (P>0.05). The expression of SCF and c-kit mRNA was positively correlated with the minimum volume threshold when AWR scored 3 and IL-10 (P<0.01), and negatively correlated with remaining indexes (P<0.01, P<0.05).@*CONCLUSION@#Moxibustion could reduce visceral hypersensitivity, improve symptoms of abdominal pain and diarrhea in IBS-D rats, and its mechanism may be related to up-regulation of the expression of SCF/c-kit signaling pathway and improvement of IBS-D immune function.
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Rats , Animals , Irritable Bowel Syndrome/therapy , Moxibustion/methods , Rats, Sprague-Dawley , Interleukin-10 , Interleukin-8 , Maternal Deprivation , Tumor Necrosis Factor-alpha , Diarrhea , Signal Transduction , Homeostasis , Receptor Protein-Tyrosine Kinases , Immunity , Immunoglobulin A , Immunoglobulin MABSTRACT
As an important component of solid tumors, mast cells show specific phenotypes in various tumor microenvironments. However, the precise mechanism of mast cell accumulation and the phenotypic features of thyroid cancer (TC) remain largely unknown. Here, we found that mast cells were obviously recruited to tumor tissue by TC-derived stem cell factor (SCF). With tumor progression, mast cell levels increased gradually. In addition, intratumoral mast cells expressed higher levels of the immunosuppressive molecule galectin-9, which effectively suppresses CD8+ T-cell antitumor immunity in vitro. Blocking galectin-9 on tumor-infiltrating mast cells reversed the immunosuppression of CD8+ T cells. In conclusion, our data elucidated novel protumorigenic and immunosuppressive roles of mast cells in TC. In addition, our results indicated that blocking mast cells may impede tumor progression and ameliorate the prognosis of TC patients.
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Objetive: To investigate the long-term expression of the piggyback (PB) transposon system expressing human interleukin-6 (IL-6), nterleukin-3 (IL-3), interleukin-15 (IL-15), stem cell factor (SCF) and granulocyte-macrophage ctimulang Factor (GM-CSF) genes in the immunodeficient mice, and to provide a simple, long-term and new method for improving the reconstruction of human immune cells in the humanized mouse models. Methods: The PB transposon plasmid (PB-GFP) containing the GFP gene was constructed, and the PB transposon plasmid (PB-5F) contaning human IL-6, IL-3, IL-15, SCF, and GM-CSF genes was constructed. The 293T cells were divided into negative control group (non-transfection), postive control group (transfected with pLVTHM), transient transfection group transfected with PB-GFP plasmid and stable transfection group transfected with PB-GFP plasmid together with transposase plasmid (super-PB)]. The proportions of GFP cells in varions groups were measured by flow cytometry every three days after transfection. The NOD. Cg-Prkdcscid IL2rgun1wj1/SzJ (NCG) mice were divided into transient transfection group and stable transfection group. The mice in transient transfection group were transfected with PB-GFP alone, and the mice in stable transfection group were transfected with PB-5F and super-PB. On the 1st, 4th, 5th and 9th days and the followng every week after the transfection, the blood samples were collected, and the serum was separed; the levels of IL-6, IL-3, IL-15, SCF, and GM-CSF in serum of the mice in various groups were detected by ELISA Results: At 30 d after transfection of PB-GFP, the percentage of GFP1 cells of the mice in stable transfection group (4. 61% + 0.42%) was significantly higher than those in postive control group (0.58% +0.05%) and transient transfection group (0.86%+ 0.10%) (P<0.05). At 30 d after transfection of PB-5F plasmid, the levels of serum IL-6, IL-15 and GM-CSF of the NCG mice in stable transfection group were significantly higher than those in transient transfection group (P
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Objective To investigate the association between stem cell factor (SCF) expression and tubulointerstitial fibrosis in the kidney of rat model with uric acid nephropathy. Methods Thirty-six Wistar rats were randomly divided into model group and control group. The rats in the model group were fed with adenine by lavage at a dose of 150 mg·kg-1·d-1, and the rats in the control group were fed with normal saline by lavage at equal volume. Six rats from each group were sacrificed respectively at week 4, 8 and 12. The mRNA levels of SCF and ColⅠin the kidney were detected by real-time fluorescence quantitative polymerase chain reaction (PCR), and their protein levels and infiltrated Mast cell (MC) were measured by immunohistochemistry. The difference and correlation of each index among different time points and groups were compared. The differences between the two groups were tested by t-test, multiple data were tested by one-way analysis of variance (ANOVA) and the correlation was analyzed by Pearson's correlation. Results ①The serum uric acid (SUA) [week 4, 8, 12: (302 ±41))μmol/L, (424 ±61) μmol/L, (518 ±57) μmol/L], creatinine[week 4, 8, 12:(151±9)μmol/L, (219±15)μmol/L, (299±21)μmol/L], urea nitrogen [week 4, 8, 12:(26.7±3.7) mmol/L, (40.3 ±5.7) mmol/L, (61.9 ±9.4) mmol/L], urine protein/creatinine (Up/Ucr) [week 4, 8, 12: (0.71 ±0.10) mmol/L, (1.18 ±0.11) mmol/L, (1.78 ±0.13) mmol/L] and the expressions of SCF mRNA [week 4, 8, 12: (1.19 ± 0.41), (1.69 ±0.63), (2.21 ±0.97)} and SCF protein [week 4, 8, 12: (1.42 ±0.33), (6.02 ±1.81), (10.03 ±2.69)] in nephridial tissue of model group's rats were significantly higher compared to the control group (all of t value>4.59, P<0.01), and the indexes were increasing gradually as the lavage going on. ②The expression of SCF was correlated with collagenⅠ, degree of renal interstitial injury, urine nitrogen, serum creatinine and UP/Ucr(At week 4, r=0.53, 0.42, 0.40 and 0.51 respectively;at week 8, r=0.60, 0.59, 0.41 and 0.39 respectively;at week 12, r=0.74, 0.61, 0.56 and 0.39 respectively, all of P value<0.05). ③ Compared with control group, the number of infiltrated MC was significantly higher in the model group, and was positively correlated with the expression of SCF (r=0.91, P<0.01). Conclusion Compared with the control group, the expression of SCF and the number of infiltrated MC in the renal interstitium are evidently increased, and are increasing gradually as the lavage going on and the deteriorating of renal interstitial damage. These results suggest that both may play important roles in the occurrence and development of uric acid nephropathy.
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The objective was to study the effect of mechanical intestinal obstruction in rats on the phenotype of interstitial cells of Cajal (ICC). Healthy Wistar rats were randomly divided into sham-operation group (C), one day obstruction group (M1), two days obstruction group (M2), and three days obstruction group (M3), with 10 rats in each group. The expression of SCF mRNA and c-Kit protein in intestinal tissue was investigated by RT-PCR and immunohistochemistry. Compared with the sham-operation group, the relative expression of SCF mRNA and the expression of c-Kit protein in intestinal tissue were significantly decreased in both obstruction groups. Levels decreased gradually with the prolongation of obstruction time, and significantly decreased on the 3rd day after obstruction (P<0.05). Immunohistochemical staining of the small intestine showed that the number of ICC in the sham-operation group was the highest, and they were gradually decreased with the extension of obstruction time in the M1 to M3 groups. There was a significant difference between groups (P<0.05). Intestinal obstruction caused a decrease in the concentrations of SCF mRNA and c-Kit protein in ICC. With the prolongation of intestinal obstruction, the number of ICCs gradually decreased.
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Animals , Male , Rats , RNA, Messenger/metabolism , Stem Cell Factor/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Interstitial Cells of Cajal/metabolism , Intestinal Obstruction/metabolism , Phenotype , Immunohistochemistry , Rats, Wistar , Disease Models, Animal , Interstitial Cells of Cajal/pathology , Intestinal Obstruction/pathologyABSTRACT
Abstract: The growth factor receptor c-kit (CD117) is expressed in immature T-cells and in some advanced forms of mycosis fungoides. c-kit gene mutation results in unrestricted neoplastic proliferation. We aimed to detect by PCR the most frequent exon mutations in seventeen plaque-stage MF patients, in their perilesional skin and in healthy skin donors. We secondarily evaluated CD117 expression by immunohistochemistry in plaque-stage and tumor-stage MF. We detected no mutation in c-kit gene and low CD117 expression was confirmed on atypical cells in one patient. Complete c-kit exon and intron sequences should be assessed and more sensitive sequencing method could be also applied.
Subject(s)
Humans , Male , Female , Aged , Exons/genetics , Mycosis Fungoides/genetics , Proto-Oncogene Proteins c-kit/genetics , Mutation/genetics , Immunohistochemistry , Case-Control Studies , Gene Expression , Polymerase Chain Reaction , Prospective StudiesABSTRACT
Skin melanin metabolism is a complex and fine regulation process, which is controlled by multiple genes, many enzymes and proteins. With the in-depth study of the factors affecting the skin melanin metabolism, scientists have found that the signaling pathway initiated by the combination of stem cell factor receptor c-kit and its ligand stem cell factor plays an important role in the metabolism of skin melanin.The paper summarizes the recent research progress on the role of c-kit receptor in the skin melanin metabolism, including the molecular mechanism and the clinical application.
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Aim To explore the change of stem cell factor-C-kit (SCF-C-kit) system in irritable bowel syn-drome with diarrhea ( IBS-D) and the correlation be-tween SCF-C-kit system and immune dysfunction . Methods Twelve neonatal rats were divided into nor-mal group and model group with six rats in each group . IBS-D rat model was established through three-factor method ( mother-son separation , acetic acid stimulation and constraint ) . Immunohistochemistry was used to observe in situ protein expressions of SCF and C-kit. qPCR was used for mRNA expressions of SCF and C-kit.Correlation between SCF-C-kit system and spleen coefficients , thymus coefficients , TNF-α, IL-8 , IL-10 was analyzed by statistics .Results Compared with normal group , positive rates of SCF and C-kit protein in model group both decreased , and so did mRNA ex-pression .Expression of SCF was negatively correlative with spleen coefficient , TNF-αexpression and IL-8 ex-pression , while positively correlative with IL-10 expres-sion.Expression of C-kit was negatively correlated with thymus coefficient , spleen coefficient and TNF-α. Conclusion SCF-C-kit system of IBS-D is abnormal, which may be related with immune dysfunction .
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Objective To explore the in vitro culture methods for oriented differentiation of peritoneal cells and bone marrow cells into high-purity mast cells,and to identify the function of these mast cells.Methods Peritoneal cells and bone marrow cells were isolated from the peritoneal cavity lavages and femur of C57BL/6 mice,and cultured with both interleukin-3 (IL-3) and stem cell factor for 2 and 4 weeks respectively.Light microscopy was performed to observe the morphology of these cells,toluidine blue staining to identify the degree of maturity of these mast cells,and flow cytometry to measure the expression of cell surface markers C D 117 and FceR Ⅰ α.After the stimulation with compound 48/80 at different concentrations,the degranulation rate of mast cells was counted under the microscope,and β-hexosaminidase release rate was measured by spectrophotometry.Results After 2-or 4-week culture,the mouse peritoneal and bone marrow cells all manifested as refractive suspension cells of uniform size.Toluidine blue staining showed violaceous metachromatic granules in the cytoplasm of the two kinds of cells.The proportions of CD117 or FcεR Ⅰ α single-positive peritoneal and bone marrow-derived mast cells were all more than 95％,and the proportions of CD117/FcεR Ⅰ α double-positive peritoneal and bone marrow-derived mast cells were 97.68％ ± 0.80％ and 96.12％ ± 0.76％ respectively.The degranulation rates of mast cells in the 100-and 1 000-mg/L compound 48/80 groups significantly differed from those in the blank control group (all P ＜ 0.01).Compared with the blank control group,the β-hexosaminidase release rates significantly increased in bone marrow-derived mast cells in the 100-mg/L compound 48/80 group and peritoneal mast cells in the 10-and 100-mg/L compound 48/80 groups (P ＜ 0.01 or 0.05).Conclusion IL-3 and stem cell factor can co-induce the directed differentiation and proliferation of mouse bone marrow stem cells and peritoneal cells,so as to harvest highnuritv mature degranulated mast cells,and lay a foundation for subsequent cell biology research.
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Objective To explore the relationship between the level of serum stem cell factor (SCF) and the activity of lupus nephritis (LN). Methods Sixty LN patients who had underwent routine renal biopsy were selected from March 2014 to September 2016. According to the systemic lupus erythematosus disease activity index (SLEDAI), the LN patients were divided into two groups: active nephritis group (30 cases) and stable nephritis group(30 cases). Meanwhile 30 healthy controls were selected as normal control group. Spearman correlation analysis test was used for correlations between the level of serum SCF and the activity of LN. Results The serum level of SCF was significantly higher in active nephritis group [(357.29 ± 63.85) ng/L] than that in stable nephritis group [(310.03 ± 40.17) ng/L] , the serum level of SCF in stable nephritis group was significantly higher than that in normal control group [(154.06 ± 22.49) ng/L], and there were significant differences (P0.05). Conclusions The level of SCF may play an important role in the occurrence and development of LN. It could reflect clinical and pathology changes and emerg as a potential serum biomarkers of LN.
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Objective@#To investigate the correlation between expression of long non-coding RNA (lncRNA) ZXF1 and clinicopathological characteristics, as well as prognosis of lung adenocarcinoma; and to explore its potential molecular mechanism.@*Methods@#A total of 83 lung adenocarcinoma tissue samples and 83 paracancerous lung tissue samples from lung adenocarcinoma patients were collected. The mRNA expression of ZXF1 in the tumors and the corresponding adjacent non-tumor tissues were determined by real-time fluorescence quantitative PCR. The correlation between ZXF1 level and clinicopathological characteristics such as age, gender, smoking history, tumor size, tumor differentiation and lymph node metastasis was evaluated. Survival analysis was performed using Kaplan-Meier and Cox multivariate regression, based on the data of a 12-56 months follow-up after surgery. In vitro ZXF1 was over-expressed in lung adenocarcinoma A549 cells, and then the proteins functionally related to ZXF1 were identified by protein array analysis.@*Results@#Of the 83 cases of lung adenocarcinoma, the ZXF1 mRNA levels in the tumor and adjacent non-tumor tissues were 8.32±3.05 and 1.05±0.47, respectively (P<0.05), and a high-level the high expression of ZXF1 in the tumor tissues was detected in 56 cases. The expression status of ZXF1 was closely correlated with the tumor differentiation and lymph node metastasis (P<0.05), but was not significantly related to age, gender and tumor size. Based on a 12-56 months follow-up, the patients with high level ZXF1 expression had a shorter disease free survival (DFS) and overall survival (OS) than that of the group with low level ZXF1 (all P<0.05). Multivariate analysis showed that ZXF1 expression, tumor size and lymph node metastasis were independent risk factors to DFS; and ZXF1 expression, tumor size, lymph node metastasis and tumor differentiation were independent risk factors to OS. The protein array data revealed that expressions of bone morphogenetic protein 5 (BMP-5)and stem cell factor receptor (SCFR)were upregulated upon overexpression of the ZXF1 in A549 cells.@*Conclusions@#lncRNA ZXF1 is overexpressed in lung adenocarcinoma, and is closely correlated with tumor differentiation and lymph node metastasis. As an independent risk factor, a high expression of ZXF1 indicates a poor prognosis for the patients. ZXF1 may influence the biological behavior of lung adenocarcinoma by enhancing the protein expression of BMP-5 and SCFR.
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Objective To investigate the effect of fractional laser on the expression of stem cell factor (SCF) and c-kit receptor in body surface of guinea pig.Methods 20 guinea pigs were adopted.The back skin of each guinea pig was divided into four zones:A zone (the skin was not treated),B zone (the skin was treated by fractional laser at a low fluence:20 mJ/cm2),C zone (the skin was treated at a medium fluence:60 mJ/cm2),and D zone (the skin was treated at a high fluence:120 mJ/cm2).The immunohistochemical method was used to detect the expression of SCF and c-kit in body surface of guinea pig.The results were quantified using an HPIAS-1000 system.Results One week later,the SCF expression level in B,C and D groups was increased obviously,which showed statistical difference compared with A group (P＜0.05);the c-kit expression level in C and D groups was increased obviously,compared with A group (P＜0.05).4 weeks later,the SCF expression level in C and D groups was higher than A and B groups;furthermore the level in D group was higher than C group (P＜0.05);the c-kit expression level in D group was higher than A and B groups (P＜0.05).Conclusions The fractional laser could generate the curative effect by increasing the expression of SCF and c-kit in body surface.
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<p><b>OBJECTIVE</b>To compare the effects of acupuncture, electroacupuncture (EA) and moxibustion on functional constipation in rats.</p><p><b>METHODS</b>Sixty male Sprague-Dawley rats were divided into a control group (=8), a model group (=11), a medication group (=8), an acupuncture group (=11), an EA group (=11) and a moxibustion group (=11) by random number table. The rats in the model group, medication group, acupuncture group, EA group and moxibustion group were treated with intragastric administration of loperamide hydrochloride for 6 days continuously to establish the functional constipation models, while equal volume of drinking water was administrated to rats in the control group at the same time. The rats in the acupuncture group, EA group and moxibustion group were respectively treated with acupuncture, EA and moxibustion at "Tianshu" (ST 25) and "Shangjuxu" (ST 37) one hour after intragastric administration; rats in the medication group were treated with intragastric administration of cisapride suspension. All the treatment was given once a day for 6 days. At the last day of intervention, the 24-hour food intake, stool quantity and its water content were measured in each group; the pushing rate of intestine was measured; the structure of colon tissue and acidic mucus in its mucous layer were observed by hematoxylin-eosin dyeing and alcian blue dyeing; the expression of stem cell factor (SCF) and c-kit mRNA was detected by real-time PCR.</p><p><b>RESULTS</b>Compared with the control group, the 24-hour food intake and stool quantity were reduced in the model group (both<0.01), and the water content of stool and pushing rate of intestine were reduced (both<0.01); compared with the model group, the stool quantity and its water content were increased in the medication group, acupuncture group, EA group (<0.05,<0.01), which were not significantly different from those in the moxibustion group (both>0.05). The pushing rate of intestine in each intervention group was increased (all<0.01). The 24-hour food intake and stool quantity in the medication group were not significantly different from those in the acupuncture group, EA group and moxibustion group (all>0.05), and the water content of stool was only reduced in the moxibustion group (<0.01). The pushing rate of intestine in the acupuncture group and moxibustion group was lower than that in the medication group (both<0.01), while that in the EA group was not significantly different from that in the medication group (>0.05). The water content of stool in the moxibustion group was lower than that in the acupuncture group and EA group (both<0.01). The pushing rate of intestine in the acupuncture group and moxibustion group was lower than that in the EA group (both<0.01). The HE staining result indicated the structure of colon tissue was normal, complete and similar in each group; the alcian blue staining indicated the acidic mucosubstance in the model group was lower than that in the control group; compared with the model group, the acidic mucosubstance in the medication group, acupuncture group, EA group and moxibustion group was all increased. Compared with the control group, the expression of SCF and c-kit mRNA was reduced in the model group (both<0.05); compared with the model group, the expression of SCF and c-kit mRNA was increased in the medication group, acupuncture group, EA group and moxibustion group (all<0.05); compared with the moxibustion group, the expression of c-kit mRNA was reduced in the acupuncture group and EA group (both<0.05).</p><p><b>CONCLUSIONS</b>Acupuncture, EA and moxibustion all can play a positive regulative role on functional constipation in rats, in which EA has the best efficacy, followed by acupuncture.</p>
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AIM:To construct AFT024-SCF cell line and HPC-Lhx2 cell line for confirming the biological function of AFT024-SCF.METHODS:The HPC-Lhx2 cell line, AFT024-SCF cell line and AFT024-GFP cell line were constructed by retro-viral infection.The expression of stem cell factor(SCF) in AFT024-SCF cells was detected by real-time PCR and Western blot.SCF in the supernatant of AFT024-SCF was detected with ELISA.The supernatant of AFT024-SCF and AFT024-GFP were collected and then diluted (1:10) with basic IMDM medium.So we made 4 culture medium:AFT024-SCF medium was used for experiment group, AFT024-GFP medium was used for endogenous negative control, IM-DM basic medium was used for exogenous negative control, and IMDM basic medium with SCF was used for positive con-trol.SCF-dependent HPC-Lhx2 cell line was cultured in these 4 different medium for 72 h.According to MTT method and colony forming experiment, the biological function of AFT024-SCF was confirmed by the proliferation ability of SCF-depend-ent HPC-Lhx2 cell line.RESULTS:SCF was highly expressed in AFT024-SCF cells.After cultured for 72 h, neither IM-DM basic medium nor GFP-AFT024 medium support HPC-Lhx2 cell line proliferation.However, AFT024-SCF medium supported HPC-Lhx2 cell line expansion as well as the positive control medium.CONCLUSION:AFT024-SCF cells ex-press SCF successfully and recombinant SCF can be replaced by the supernatant of AFT024-SCF culture medium for expan-ding HPC-Lhx2 cell line in vitro.
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Objective To explore the effect and intracellular signal transduction pathway of insulin-like growth factor 1 (IGF-1) on the expression of stem cell factor (SCF) in spesis rats colonic smooth muscle cells (SMC).Methods The model of spesis rats was established,colonic SMC from spesis rats were cultured by enzymolysis and identified by α-actin immunofluoresence methods.Colonic SMC incubated with different concentrations (0,50,100 and 150 μg/L) of IGF-1 for different durations (0,12,24,36 and 48 h),and another SMC were pretreated with specific mitogen-activated protein kinase (MEK) inhibitor PD-98059 and phosphatidylinostiol 3-kinase(PI 3-kinase) inhibitor LY-294002 before IGF-1 induced.The expression of SCF in colonic SMC was examined by Western blot and quantitative reverse transcription polymerase chain reaction.Results (1) A very low level of SCF was expressed in colonic SMC cultured in bovine serum free medium.The protein and mRNA were 0.223 ± 0.047 and 0.149 ± 0.023 respectively.The expression of SCF protein and mRNA from SMC was increased to 0.398±0.033 and 0.243±0.013(P＜0.05) when concentration of IGF-1 was 50 μg/L.The final effective concentration of IGF-1 in vitro was 100 μg/L,the levels of SCF protein and mRNA were increased to 0.719±0.017 and 0.360±0.006(P＜0.05).(2) For the time point induced the SCF expression by IGF-1,the expression level of protein and mRNA in 0 h were 0.347±0.031 and 0.092 ±0.018 respectively.The time point of SCF peak level occurred at the 24 h,the expression level of SCF protein and mRNA were increased to 0.893 ± 0.049 and 0.396 ± 0.048 (P ＜ 0.05).(3) The IGF-1-induced SCF expression was reduced significantly by a pretreatment of PD-98059 (SCF protein and mRNA were reduced 37％ and 45％,P＜0.05),and LY-294002 had no effect on the expression of SCF (P＞0.05).Conclusion The expression of SCF from colon SMC in sepsis rat is stimulated by IGF-1 in both dose and time-dependent manners,in which ERKMAPK signal transduction may play an important role.
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Objective To explore the effect of stem cell factor (SCF), Flt3 Ligand (FL) and Thrombopoietin (TPO) on the expansion of the mononuclear cells derived from umbilical cord blood ex vivo under hypoxic conditions.Method Human fresh CB mononuclear cells were isolated and cultured with different combinations of cytokine for 7 days under normoxic or hypoxic conditions (3 ％ O2).At day 0 and 7 of culture, total nucleated cells (TNC), CD34+ , CD34+ CXCR4+ , CD34+ CD49d+ and CD34+ CD62L+ cells were assayed by flow cytometry, and colony forming unit (CFU) was determined.The experiment was divided into four groups: group A (cells were cultured in single mononuclear cells under normoxic conditions);group B (cells were cultured in single mononuclear cells under hypoxic conditions);group C (cells were cultured with SCF, FL and TPO under normoxic conditions);group D (cells were cultured with SCF, FL and TPO under hypoxic conditions).Result At 7th day of culture, TNC, CD34+ cells and CFU expansion were significantly increased in groups B, C and D, and the expansion effect in group D was superior to that in groups B and C, but there was no significant difference between groups B and C (P＞0.05).At 7th day of culture, the expression levels of CD49d, CD62L and CXCR4 on expanded CD34 + cells were significantly reduced in group A as compared with those at day 0, and those in groups B, C and D were significantly increased as compared with those in group A, those in group D were significantly higher than in group D than in groups B and C (P＜0.05),but there was no significant difference between groups B and C (P＞ 0.05).Conclusion Hypoxic conditions could enhance the expansion and expression of CD49d, CD62L and CXCR4 of mononuclear cells derived from umbilical cord blood with SCF, FL and TPO.
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Objective To study the effects of Bacillus Calmette-Guerin (BCG) intervention on the expression of stem cell factor (SCF) in asthma. Methods Kunming mice were randomly divided into asthmatic group, BCG group and control group of ten mice each group. Mice were sensitized and challenged with ovalbumin (OVA) to establish asthmatic model. After twenty-four hours of last challenge, eosinophils (EOS) were counted in the lung tissue and bronchoaveolar lavage fluid (BALF). The level of SCF in BALF was determined by enzyme-linked immunosorbent assay. The expression of SCF protein in lung tis-sue was measured by immunohistochemistry technique and computerized image analysis system. Results The number of EOS in lung tissue was (10.67±1.94)/HP, (6.40±1.55)/HP and (0.37±0.33)/HP in asthmatic, BCG and control group respectively. The number of EOS in BALF was (7.58 ± 1.30) × 107/L, (3.78 ± 1.15) × 107/L and 0 in asthmatic, BCG and control group respectively. The difference between each group was statistically significant (P0.05). In asth-matic group, the expression of SCF in lung was positively correlated with the number of EOS (P<0.05). Conclusions BCG treat-ment can markedly decrease the airway inflammation in asthmatic mice. BCG cannot inhibit the expression of SCF in asthma.
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AIM: To investigate the expression of stem cell factor (SCF) in tryptase -positive mast cells ( MCs) in different types of human periapical diseases for determining the role of SCF and MCs in the pathogenesis of peria-pical diseases.METHODS: A total 50 cases of specimens were involved in this study, including healthy control (n=20), periapical cyst (n=15) and periapical granuloma (n=15).The tissue material was fixed in 10%formalin for at least 48 h, stained with hematoxylin and eosin for the observation of histopathology, stained with immunohistochemistry for identifying MCs and MCs degranulation, and stained with double immunofluorescence for identification of tryptase-SCF double positive MCs.RESULTS:Compared with healthy control, significantly higher densities of both total and degranu-lated MCs in human periapical lesions were observed.The densities of both total and degranulated MCs in the periapical cyst were significantly higher than that in the periapical granuloma.The density of tryptase-SCF double positive MCs in the periapical lesions was significantly higher than that in the healthy controls.The density oftryptase-SCF double positive MCs in the periapical cyst was significantly higher than that in periapical granuloma.No significant difference in the density of MCs between immunohistochemistry staining and double immunofluorescence staining was observed.CONCLUSION:The tryptase-SCF double positive MCs play an active role in the pathogenesis of the periapical inflammatory lesions, particularly in the formation of fibrous tissue in periapical cyst.The potential role of the tryptase-SCF double positive MCs relates with the initiation, development, and persistence of the periapical inflammatory process.