Induction Of Helicobacter Pylori L Form In Vitro; The Infection Status Of Helicobacter Pylori Among Patients And The Relationship With Clinicopathologic Features In The Middle Area Of Hexi Corridor

Objective:1. Induction of Helicobacter pylori (Hp) L form in vitro with different concentration of penicillin in order to further explore the biological characteristics, the mechanism of L form formation and the effect of Hp-L form in the people suffered with upper gastrointestinal tract disease.2. To explore the status of Hp infection among people suffered with upper gastrointestinal tract disease in the middle area of Hexi Corridor and analyze the relationship between Hp infection and clinicopathologic features.3. To study the expression of Cyclin D1 in upper gastrointestinal diseases of different pathologic tissues and the relationship of the Cyclin D1 expression and Hp infection in the mucosa of stomach.Methods:1.Certain concentration and volume of Hp were spotted onto L-form induction media(LIM) which contained different concentration of penicillin at 2400ug/ml,1200ug/ml,600ug/ml,300ug/ml,150ug/ml and 75ug/ml respectively and then incubated at microaerophilic air condition at 37℃about 5 to 7 days. The morphology of the bacterial colony was observed in the inverted microscope and the Gram's staining were used to observe the bacterial forms.2. The samples of gastric mucosa embedded in paraffin and pathologic data of the patients during the time from 2003 to 2005 were collected from the People's Hospital of Zhangye Municipality. Warthin-Starry silver staining and Gram's staining were used to detect Hp infection of the samples.the relationship between Hp infection and clinicopathologic features was analyzed.3.Tissue chip were made and immunohistochemical stain was used to detecting the expression of Cyclin D1 in the mucosa of stomach.The relationship of the Cyclin D1 expression and Hp infection was analyzed.Results:1. After 5 days microaerophilic incubation at 37℃, lots of typical "fried egg" L-form colonies were appearing in the LIM contained 1200ug/ml and 600ug/ml penicillin which were observed in the inverted microscope.The bottom of the L-form colonies embedded into the media. Most morphology of the bacteria in the "fried egg" L-form colonies were negative coccoid bacteria after Gram's staining. LIM contained 2400ug/ml,300ug/ml,150ug/ml, and 75ug/ml penicillin can only see a few L-form colonies with the same morphology as negative coccoid bacteria.2. The positive rate of Hp infection was 81.3%(178/219) in the 219 samples. The positive rate of Hp infection in gastric cancer group, precancerous lesion group and superficial gastritis group were 85.6%,77.4%, and 60.7% respectively. There exist significant difference among the three groups (P<0.05). There exist significant difference of Hp infection rate between gastric cancer group and superficial gastritis group (P<0.05). Hp infection rate among people over age of 61 was significantly higher than that under 60 (P<0.05). Hp infection in Gastric cancer with lymph node metastasis was significantly higher than that without lymph node metastasis (P<0.05). There was no significant difference of Hp infection among sex and different gastric cancer pathologic forms, cancer sizes and invasion (P>0.05)3. The positive rate of Cyclin D1 expression among gastric cancer group, precancerous lesion group and superficial gastritis group were 58.7%,56.7%, and 18.5% respectively. The positive rate of Cyclin D1 expression was significantly higher in gastric cancer group and precancerous lesion than that in superficial gastritis group. The positive rate of Cyclin D1 expression in Gastric cancer with lymph node metastasis was significantly higher than that without lymph node metastasis (P<0.05). There exist significant difference of Cyclin D1 expression in different invasion of gastric mucosa (P<0.05).4. There was no significant difference of the positive rate of Cyclin D1 expression between the Hp infected group (52.7%) and non-infected group (54.3%) (P>0.05).Conclusion:1. Appropriate levels of penicillin (1200ug/ml or 600ug/ml) can induce a large number of Hp L form formation in vitro. 2. Hp infection among people suffered with upper gastrointestinal tract disease in the middle area of Hexi Corridor was 81.3%. Hp infection involves in the development of upper gastrointestinal tract disease and is increased with the age in the area and it is related to lymph node metastasis of gastic cancer.3. Cyclin D1 expression involves in the development of gastric cancer and is related to lymph node metastasis and invasion depth. These indicate that Cyclin D1 is an important mark of the gastric cancer prognosis.4. There was no relationship between the expression of Cyclin D1 and Hp infection in the mucosa of stomach.