Efforts have been made to accomplish a long term in vitro cultivation of mouse peritoneal macrophage as a host cell for growth of Mycobacterium lepraemurium. Following the inoculation with live or heat-killed Myco. lepraemuriuum of cultured macrophages either on a cover-slip in Leighton tube or in a small petri dish, microscopic observations of acid-fast (AF) stained slide preparation, and total counts of AF bacilli that were released by ultrasonic treatment from the macrophages in small petri dish have been followed in order to present microscopic and quantitative evidence of the actual multiplication of Myco. lepraemurium in cultured mouse peritoneal macrophage. The results are summarized and conclusions are as follows; 1. Successful long term in vitro cultivation of mouse peritoneal macrophage has been accomplished. The growth medium for tissue culture consisted of NCTC 109;50% heat-inactivated calf serum; 40% and beef embryo extract (diluted 1 : 5); 10% and the medium was renewed every 3 to 4 days. The incubation temperature was 37 degrees C; before and at 30 degrees C; after the inoculation with Myco. lepramurium. The CO2 content inside the CO2 humidity incubator for the cultivation of macrophage was kept at 5%. 2. In cultures of macrophage inoculated with live Myco. lepraemurium, clear features of increases in the number of AF bacilli inside individual cell, of elongation of bacill and of increased solidity in AF staining were observed. However, these features were absent in cultlires of macrophage inoculated with heat-killed Myco. lepraemurium. 3. The ultrasonic treatment of macrophage inoculated with 1ive Myco. lepraemurium, and the quantitative assessment of total number of AF bacilli through the course of 6 to 8 weeks after inoculation has provided partial but substantial evidence of actual multiplication of Myco. lepraemurium in cultured macrophages.
Animal ; Female ; Macrophages/microbiology* ; Mice ; Mycobacterium leprae/growth & development* ; Peritoneum/cytology ; Tissue Culture

OBJECTIVETo study the effects of advanced glycosylation end products (AGEs) on the production of fibronectin (FN) in human peritoneal mesothelial cells (HPMC) in vitro and the role of protein kinase C (PKC) in this course.

METHODSThe AGE-human serum albumin (HSA) (0, 100, 500, 1 000 microg/ml) was used in culture medium to stimulate the HPMC. The mRNA level of FN was measured with real-time PCR, moreover, the protein level of FN in HPMC was detected by ELISA. With the method of ELISA, the PKC activities were observed. Inhibitors or activators of PKC were used to observe the roles of PKC pathways on the AGE-HSA stimulated productions of FN in HPMC.

RESULTSAGE-HSA activated PKC in HPMC in a dose, time-dependent manner (P < 0.05). AGE-HSA up-regulated the expression of FN mRAN and protein in dose- and time-dependently (P < 0.01); PKC activator phorbol 12-myristate 13-acetate (PMA) induced FN expression, respectively depletion of PKC and calphostin C, a PKC inhibitor, effectively prevented both PMA and AGE-HSA-induced expression of the FN (P < 0.05).

CONCLUSIONAGEs can increase the activities of PKC. AGEs can directly increase FN expression in HPMC which may contribute to peritoneal fibrosis and this is regulated by PKC.

Cells, Cultured ; Epithelium ; secretion ; Fibronectins ; metabolism ; Glycation End Products, Advanced ; pharmacology ; Humans ; Peritoneum ; cytology ; Protein Kinase C ; metabolism

OBJECTIVETo investigate the effect of Changtong oral liquid (CTOL) on the proliferation of cultured fibroblasts derived from normal peritoneum (NFs) and adhesive peritoneum (AFs) of rats.

METHODSTwenty male SD rats were randomized into 4 groups, including a normal serum group and 3 CTOL groups with CTOL treatment at low, medium or high doses. Serum samples were obtained from the abdominal arteries of the rats after oral treatment with CTOL for 7 days. The fibroblasts were isolated from the peritoneum by means of tissue culture, and the passage 3-8 cells were cultured with the sera of the normal control and CTOL groups for 24, 48, 72 and 96 h. MTT assay was used to observe the proliferation of the fibroblasts.

RESULTSThe dose of CTOL was inversely correlated to the absorbance but positively to the growth inhibition rates. Compared with the NFs cultured in normal control rat serum, the NFs in serum from CTOL groups showed no obviously changes in the absorbance at 24 and 48 h, but displayed significant reduction at 72 and 96 h (P<0.01). Compared with the AFs in normal rat serum, the AFs in the 3 CTOL groups all showed significantly decreased absorbance at 24, 48, 72 and 96 h (P<0.05). At the same time point, the inhibition rate of AFs in low-dose CTOL group showed no significant difference from that in the normal control group, but CTOL at a medium dose resulted in a significantly higher inhibition rate of AFs at 72 h (P<0.05). High-dose CTOL produced significant differences in the inhibition rates of AFs and NFs (P<0.05).

CONCLUSIONCTOL can inhibit the proliferation of AFs and NFs in vitro. AFs appear to be more sensitive to CTOL, which has a dose-dependent inhibitory effect of AF proliferation.

Adhesiveness ; Administration, Oral ; Animals ; Cell Proliferation ; drug effects ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; administration & dosage ; pharmacology ; Fibroblasts ; cytology ; drug effects ; Male ; Peritoneum ; cytology ; Rats

OBJECTIVE: To investigate the role of phosphorylation of glycogen synthase kinase-3beta (GSK-3beta) inducing epithelial mesenchymal transition in human peritoneal mesothelial cells (HPMC). METHODS: Primary HPMC was harvested from human omental tissue and maintained under defined in vitro conditions. The expression of p-GSK-3beta and total GSK-3beta in HMPC was detected by Western blot after incubation with different concentrations (0, 5, 10, 20, and 40 mmol/L)of LiCl at different time points (0, 1, 3, 6, and 12 h). The protein expression of E-cadherin and alpha-SMA was also examined after treatment with 20 mmol/L LiCl according to different time courses. The intracellular distribution and expression of alpha-SMA were determined by indirect immunofluorescence. RESULTS: LiCl stimulated phosphorylation of GSK-3beta and the effect was time-dependent and concentration-dependent to limited extent (P<0.05). The expression of alpha-SMA increased (P<0.05) and the expression of E-cadherin decreased significantly (P<0.05) after 24 h stimulation by 20 mmol/L LiCl. The indirect immunoflurescence showed that the expression of alpha-SMA in HPMC increased significantly after 24 h incubation with 20 mmol/L LiCl. CONCLUSION The phosphorylation of GSK-3beta leads HMPC to epithelial mesenchymal transition and provides new clue for the treatment of peritoneal fibrosis.
Actins ; metabolism ; Cadherins ; metabolism ; Epithelial Cells ; cytology ; drug effects ; Epithelial-Mesenchymal Transition ; drug effects ; Glycogen Synthase Kinase 3 ; metabolism ; Glycogen Synthase Kinase 3 beta ; Humans ; Lithium Chloride ; pharmacology ; Mesoderm ; cytology ; drug effects ; Peritoneum ; cytology ; Phosphorylation

OBJECTIVE: To determine the effect of indomethacin on high concentration glucose and lipopolysaccharide (LPS) induced fibronectin (FN) and plasminogen activator inhibitor-1 (PAI-1) secretion in cultured human peritoneal mesothelial cells. METHODS: Mesothelial cells were isolated from human omental specimens by trypsin disaggregation and incubated by 2.5% glucose or LPS together with different concentrations of indomethacin. Enzyme-linked immunosorbent assay determined the quantity of FN and PAI-1 in the cultured supernatants. RESULTS: Compared with the control group, the levels of FN and PAI-1 in the cultured supernatants were increased significantly exposuring to high concentration glucose and LPS (P <0.01). The different concentrations of indomethacin decreased FN and PAI-1 secretion in the 2.5% glucose or the LPS group within 24 h (P < 0.05). CONCLUSION Indomethacin can inhibit the synthesis and secretion of extracellular matrix in human peritoneal mesothelial cells, which may be effective in the gene therapy for peritoneal fibrosis.
Cells, Cultured ; Cyclooxygenase Inhibitors ; pharmacology ; Epithelial Cells ; metabolism ; Fibronectins ; metabolism ; Humans ; Indomethacin ; pharmacology ; Peritoneum ; cytology ; metabolism ; Plasminogen Activator Inhibitor 1 ; metabolism

OBJECTIVE: To investigate the possible role of Danshensu on fibronectin (FN) and collagen-I (Col-I) secretion induced by high glucose in human peritoneal mesothelial cells (HPMCs). METHODS: HPMCs were treated with high glucose and Danshensu at different concentrations. The mRNA expression of FN, Col-I, endothelin-1 (ET-1), and heme oxygenase-1 (HO-1) were examined by reverse transcription-polymerase chain reaction (RT-PCR). The protein expression of FN, Col-I HO-1 and ET-1 were analyzed by enzyme-linked immunosorbent assay or immunofluorescence method. RESULTS: The expression of protein and mRNA of FN and Col-I were attenuated by Danshensu in both dose-dependent and time-dependent manner. The mRNA and protein levels of ET-1 were decreased, and the mRNA and protein levels of HO-1 increased in the Danshensu groups in a dose-dependent manner compared with the high glucose group. The expression of ET-1 and HO-1 showed little difference in a time gradient of danshensu(P>0.05). CONCLUSION Danshensu can protect HPMCs through inhibiting the expression of FN and Col-I induced by high glucose, which is related to the suppression of oxidative stress.
Cell Line ; Cells, Cultured ; Collagen Type I ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Epithelial Cells ; metabolism ; Fibronectins ; metabolism ; Glucose ; pharmacology ; Humans ; Lactates ; pharmacology ; Peritoneum ; cytology

OBJECTIVETo test the immunity of peritoneal monocytes against Plasmodium yoelii infected red blood cells (target cells).

METHODSSaponinized Plasmodium yoelii infected red blood cells (SPRBC, Ghost erythrocyte) were used to immunize mice i.p twice. Three weeks later, the infected red blood cells were injected i.p.; 90 min later, the total peritoneal cells were isolated and washed for scanning electromicroscopy to observe the effects of the peritoneal monocyte to the target cell.

RESULTSThe peritoneal cells of the immunized mice were activated after 90 min of the challenge of target cells. The size of the cell was not even and the pili on the cell surface turned to be long and densed. Cell interconnections were found among the cells. In some peritoneal monocytes, their cell plasma were scattered (omlette-like) or with the shape as "cellular bomb". The scattered or the sheeted pili and spredding cell plasma could adhere to the target cells which were perforated densely and damaged.

CONCLUSIONThe protective adaptive immunity exists in the peritoneal monocytes of immunized mice.

Animals ; Antibodies, Protozoan ; immunology ; Erythrocyte Membrane ; parasitology ; Female ; Malaria Vaccines ; immunology ; Mice ; Mice, Inbred BALB C ; Monocytes ; immunology ; ultrastructure ; Peritoneum ; cytology ; Plasmodium yoelii ; immunology ; ultrastructure

OBJECTIVETo investigate the changes of the expression of intercellular adhesion molecule-1 (ICAM-1) and integrin beta(1) in peritoneal mesothelial cells during laparoscopy-assisted radical gastrectomy(LARG) and to explore the possible effects of LARG on the peritoneal metastasis.

METHODSFrom April to August 2008, LARG was performed for 26 patients with gastric cancer (laparoscopy group), while 20 cases underwent open radical gastrectomy(open group). Peritoneum of right upper belly was collected at 3 operation time points(the beginning, 2 hours, 4 hours). The expressions of ICAM-1 and integrin beta(1) in peritoneal mesothelial cells at 3 time points were detected by immunohistochemistry.

RESULTSWith the operation prolonging, the expression of ICAM-1 and integrin beta(1) was increased gradually in both LARG and open groups. The expression of integrin beta(1) in two groups was obviously increased at 4-hour time point as compared to the beginning(P<0.05). Besides, there were no significant differences of these two adhesion molecules among the three operation time points between two groups(P>0.05).

CONCLUSIONSCompared with open surgery, LARG is not associated with a greater effect on the expression of ICAM-1 and integrin beta(1) in peritoneal mesothelial cells, and may not promote peritoneal metastasis of gastric cancer through increasing the expression of adhesion molecule in peritoneal mesothelial cells.

Adult ; Aged ; Epithelial Cells ; metabolism ; Female ; Gastrectomy ; methods ; Humans ; Integrin beta1 ; metabolism ; Intercellular Adhesion Molecule-1 ; metabolism ; Laparoscopy ; Male ; Middle Aged ; Peritoneum ; cytology ; Stomach Neoplasms ; metabolism ; pathology ; surgery

OBJECTIVE: To investigate the expression of galectin-1 with the stimulation of peritoneal dialysis solution (PDS) and its role in the epithelial-to-mesenchymal transition (EMT) in human peritoneal mesothelial cells (HPMCs). METHODS: HPMCs were stimulated with PDS containing different concentrations of high glucose (1.5%, 2.5%, and 4.25%). After 24 h, mRNA and protein expressions of galectin-1,vimentin, and zo-1 were analyzed with real-time PCR and Western blot, respectively. Liposome transfected siRNA technique was used to knock down the expression of galectin-1 and the effect of galectin-1 siRNA on the EMT of HPMCs was also observed under 4.25% PDS condition. RESULTS: mRNA expression of galectin-1 in HPMCs increased in PDS groups, especially in group with 4.25% PDS (P<0.05). Protein expression of galectin-1 in HPMCs significantly increased in PDS groups with a dose dependent manner (P<0.05).Expression of vimentin in HPMCs significantly increased in PDS groups, especially in groups of 2.5% PDS and 4.25% PDS (P<0.05), but zo-1 expression markedly decreased (P<0.05). The expression of galectin-1 correlated positively with vimentin (P<0.05) but negatively with zo-1 (P<0.05). Expression of vimentin in groups of 4.25% PDS was markedly inhibited (P<0.05) by galectin-1 siRNA, whereas zo-1 expression was significantly increased (P<0.05). CONCLUSION Galectin-1 can mediate high glucose PDS-induced EMT in HPMCs and may be a new target for the prevention and treatment of peritoneal fibrosis.
Cells, Cultured ; Dialysis Solutions ; pharmacology ; Epithelial Cells ; cytology ; Epithelial-Mesenchymal Transition ; drug effects ; Galectin 1 ; genetics ; metabolism ; Glucose ; pharmacology ; Humans ; Peritoneal Dialysis ; adverse effects ; Peritoneal Fibrosis ; etiology ; Peritoneum ; cytology ; RNA, Messenger ; genetics ; metabolism

INTRODUCTIONPeritoneal washing cytology and imprint cytology of pelvic lymph nodes samples were used to evaluate the rapid cytologic detection of peritoneal and retroperitoneal spread of endometrial cancer.

MATERIALS AND METHODSWe undertook a study on 194 endometrial cancer patients who underwent primary treatment in the Gynecologic Clinic, Democritus University of Thrace. All patients were subjected to peritoneal washing (PW) cytology and imprint cytology performed on lymph node sampling. The cytologic specimens were stained by May-Grünwald Giemsa (MGG) and Haematoxylin eosin (HE) techniques. Cell-blocks prepared from peritoneal washings (PWs) and the lymph node samples were sent for histologic examination. The cytologic fi ndings were correlated to histologic results.

RESULTSRapid intraoperative cytology provides a useful diagnostic technique for the assessment of endometrial cancer spread. HE and MGG stain presented different values of sensitivity and specifi city in the detection of peritoneal and retroperitoneal spread of endometrial cancer.

CONCLUSIONCytologic assessment of intraperitoneal and retroperitoneal spread of endometrial cancer is a rapid, intraoperative procedure, which provides the surgeon with useful information regarding the stage of the disease and the subsequent therapeutic approach.

Cytodiagnosis ; Endometrial Neoplasms ; diagnosis ; pathology ; Eosine Yellowish-(YS) ; Female ; Greece ; Humans ; Intraoperative Period ; Lymph Nodes ; cytology ; pathology ; Methylene Blue ; Peritoneal Neoplasms ; diagnosis ; pathology ; secondary ; Peritoneum ; cytology ; pathology ; Time Factors