Study of PIK3CA, BRAF, and KRAS mutations in breast carcinomas among Chinese women in Qinghai.

Phosphatidylinositol-3-OH kinase and RAS-activated signaling pathways play an important role in tumor formation. Abnormalities in relevant genes play essential roles in the occurrence and development of many human cancers. Studies of breast cancer have mainly focused on the women in western countries, but few studies have examined the frequency of mutations in PIK3CA, BRAF, and KRAS in Chinese breast cancer patients. In this study, we conducted sequence analysis of PIK3CA, BRAF, and KRAS and determined relationships with the occurrence of breast cancer in women from Qinghai. DNA was extracted from 25 cases of human breast cancer tissue samples. PIK3CA, BRAF, and KRAS mutation analysis was performed by polymerase chain reaction and DNA sequencing. No mutations were found in PIK3CA, BRAF, and KRAS of adjacent tissues. However, PIK3CA mutations were observed in 32% (8) of the 25 breast cancer tissues examined, in which exon 9 accounted for 4% (1), exon 20 accounted for 28% (7), and no mutations were found in exon 1 of PIK3CA. Sequencing of exon 2 of KRAS suggested that 20% (5) of the 25 samples harbored a mutation and 16% (4) of BRAF harbored a mutation. Any mutation in these 3 oncogenes may induce the occurrence and development of breast cancer.

Effect of phosphorylation of MAPK and Stat3 and expression of c-fos and c-jun proteins on hepatocarcinogenesis and their clinical significance.

AIM: To study the effect of phosphorylation of MAPK and Stat3 and the expression of c-fos and c-jun proteins on hepatocellular carcinogenesis and their clinical significance. METHODS: SP immunohistochemistry was used to detect the expression of p42/44(MAPK), p-Stat3, c-fos and c-jun proteins in 55 hepatocellular carcinomas (HCC) and their surrounding liver tissues. RESULTS: The positive rates and expression levels of p42/44(MAPK), p-Stat3, c-fos and c-jun proteins in HCCs were significantly higher than those in pericarcinomatous liver tissues (PCLT). A positive correlation was observed between the expression of p42/44(MAPK) and c-fos proteins,and between p-Stat3 and c-jun but there was no significant correlation between p42/44(MAPK) and p-Stat3 in HCCs and their surrounding liver tissues. CONCLUSION: The abnormalities of Ras/Raf/MAPK and JAKs/Stat3 cascade reaction may contribute to malignant transformation of hepatocytes. Hepatocytes which are positive for p42/44(MAPK), c-fos or c-jun proteins may be potential malignant pre-cancerous cells. Activation of MAPK and Stat3 proteins may be an early event in hepatocellular carcinogenesis.

A further study on capillary-like bile ductules proliferation in chronic active hepatitis.

64 cases of chronic active hepatitis (CAH) and 84 cases of non-CAH liver diseases were studied with keratin stain. The capillary-like bile ductules (CLBD) were proliferating and their morphology was identical to that with Type V collagen stain reported before. CLBD proliferation were more marked in CAH than in other liver diseases, and it was considered to be one of the characteristics of CAH and could be used for differential diagnosis. The ultrastructure of CLBD was specific in morphology. The HBV-DNA in CLBD shown by the technique of in situ hybridization suggested that HBV might infect the cells of CLBD.

Increase of calcium levels at interphase in NIH 3T3 cells.

The fluorescent calcium ion indicator dye Fluo-3 and DNA-binding dye Hoechst 33342 were employed to determine, in a quantitative microspectrofluorometric study, the intracellular calcium ion concentration ([Ca2+]i) and the DNA content of individual living NIH3T3 cells. The well-separated excitation and emission properties of these dyes allowed us to establish for each cell both the phase of the cell cycle using DNA content and [Ca2+]i. We found that the transition from G1, through S, to the G2 phase is accompanied by a two-fold increase in [Ca2+]i. The [Ca2+]i was inhomologous in each phase of the interphase (G1, S and G2) although [Ca2+]i in the S and G2 phases was never lower than certain threshold values in the G1 and S phases respectively. [Ca2+]i in G0 cells was lower than that in G1 cells. These changes in [Ca2+]i suggest that [Ca2+]i may be an important regulator of cell cycle progression.

A comparison between endoscopic trans-sphenoidal surgery and traditional trans-sphenoidal microsurgery for functioning pituitary adenomas.

This retrospective study reviewed and compared the efficacy and safety outcomes following trans-sphenoidal endoscopy or microsurgery approaches in patients with functioning pituitary adenomas: 68 patients underwent endoscopic trans-sphenoidal resections and 59 patients had microsurgical resections. Tumours were classified according to diameter and clinical outcomes were evaluated. Overall disease control rates were 70.6% following endoscopy and 49.2% following microsurgery. The most obvious between-group difference was observed in macroadenomas: disease control rates were 63.9% following endoscopy and 27.3% following microsurgery. Cerebrospinal fluid leaks, diabetes insipidus and syndrome of inappropriate antidiuretic hormone secretion were observed postoperatively in both groups. The complication rate was lower following endoscopy compared with microsurgery (this difference was not statistically significant). Trans-sphenoidal endoscopy resection achieved good results in pituitary tumours, particularly for the complete removal of macroadenomas, and was an effective alternative to microsurgery.

[Role of functional inactivation of p53 from MDM2 overexpression in hepatocarcinogenesis].

OBJECTIVE: This paper was to evaluate the role of p53 mutation, and p53 and MDM2 proteins expression in hepatocarcinogenesis. METHODS: Using streptavidin-peroxidase conjugation method (SP), the expression of p53 and MDM2 proteins was observed in 61 cases of primary hepatocarcinomas (HCC) and 59 cases of corresponding paracancerous tissue, among which p53 mutations in exons 5-8 were detected in 21 cases by polymerase chain reaction single-strand confirmation polymorphism analysis (PCR-SSCP). RESULTS: Positive nuclear p53 and MDM2 immunostainings were demonstrated in 57.38% (35/61) and 26.23% (16/61) of HCC, and 1.69% (1/59) and 3.39% (2/59) of corresponding paracancerous tissue, respectively. The expressions of p53 and MDM2 proteins in HCCs were significantly higher than those in paracancerous tissues (P < 0.01). The expressions of p53 and MDM2 were not significantly correlated (P > 0.05). There were 42.86% (9/21) mutations in exon 7 of p53 gene and no mutation was observed in exons 5, 6, 8 in HCCs and in paracancerous tissues. In cases of p53 mutations, there were 66.67% (6/9) of p53 overexpression and 11.11% (1/9) of overexpression of both p53 and MDM2. MDM2 overexpression also appeared in 25% (3/12) of cases without mutation. CONCLUSIONS: Mutation of p53 gene and functional inactivation of p53 resulting from MDM2 overexpression play an important role in carcinogenesis of HCC. It is possible that p53 mutations and MDM2 overexpression induced by other mechanisms are involved in carcinogenesis of HCC.

[Effect of hepatitis C virus NS3 protein on expression of iNOS mRNA in hepatocarcinogenesis].

OBJECTIVE: To investigate hepatocarcinogenesis by detecting the effect of HCV NS3 protein on expression of inducible nitric oxide synthase (iNOS) mRNA in hepatocellular carcinoma (HCC) and pericarcinomatous liver tissues(PCLT). METHODS: The expression of HCV NS3 protein and iNOSmRNA was detected by immunohistochemical technique (SP method) and in situ hybridization in specimens of HCC and PCLT from 52 patients with HBV(-). The relationship between the positive rate and the signal strength was tested statistically. RESULTS: The positive rate of HCV NS3 protein in HCC was lower (46.2% vs. 67.3%), and signal strength was weaker than those in PCLT. The signal strength of HCV NS3 protein was correlated with the degree of carcinomatous cells differentiation in HCC (P < 0.01). The positive rates (80.8% and 90.4%) of iNOSmRNA in HCC and PCLT had no statistical difference. Signal intensity of iNOSmRNA in HCC was weaker than that in PCLT. The higher HCVNS3 protein expression level was, the stronger iNOSmRNA signals in PCLT (P < 0.01). In particular, the localization and patterns of HCV NS3 protein were much similar to those of iNOSmRNA in the PCLT of some patients. CONCLUSIONS: HCV NS3 protein may exert its hepatocarcinogenic effect in the early stage on host cells by increasing product of nitric oxide, which may bring about transformation of hepatocytes.

[Expressions of p-MAPK, cyclin D1, p53 protein and their relationship in osteosarcoma].

OBJECTIVE: To detect the expressions of p-MAPK, cyclin D1 and p53 protein, and investigate their relationship in osteosarcomas. METHODS: SP immunohistochemical technique was used to detect the expressions of of p-MAPK, cyclin D1 and p53 protein among the 58 osteosarcomas and 14 osteoid osteomas samples. RESULTS: The positive rates of p-MAPK, cyclin D1 and p53 protein were 77.6% (45/58), 60.3% (35/58) and 44.8% (26/58) respectively in 58 osteosarcomas, and 21.4% (3/14), 21.4% (3/14) and 7.1% (1/14) in 14 osteoid osteomas. Their positive rates and expressive intensities in osteosarcomas were higher than those in osteiod osteomas. Their positive correlation was significantly observed between p-MAPK and cyclin D1 proteins (P < 0.01), and wasn't observed between p-MAPK and p53 proteins (P < 0.05) in osteosarcomas. CONCLUSIONS: MAPK phosporylation may be one of causes for activation of cyclin D1, which play an important role in osteosarcogenesis. The mutation of p53 oncogene may not result in osteosarcogenesis by MAPK signaling pathway. Detecting p-MAPK by immunohistochemistry may be one of diagnostic indexes for osteosarcoma and benign osteoma.

[p16 and cyclinD1 protein expression and p16 gene mutation in primary human hepatocellular carcinoma].

OBJECTIVE: To elucidate the relationship between expression of p16 and cyclinD1 proteins and evaluate the role of p16 gene exon 2 mutation in hepatocellular carcinogenesis. METHODS: Streptavidin-peroxidase conjugated method(SP) was used to detect expressions of p16 and cyclinD1 proteins in 44 cases of hepatocellular carcinoma(HCC), and polymerase chain reaction single-strand conformation polymorphism analysis(PCR-SSCP) to detect p16 gene mutation in exon 2 in 12 cases of HCC and liver tissues adjacent to HCC. RESULTS: Expression rate and positive signal intensity of p16 protein in HCC were significantly lower(P < 0.01) and those of cyclinD1 protein in HCC were significantly higher (P < 0.05) than those in pericarcinomatous tissues. Of 12 fresh HCC tissues, p16 gene mutation in exon 2 was found in 2 cases, whereas that was not found in pericarcinomatous tissues. CONCLUSIONS: 1. Inactivation or/ and deletion of p16 protein may be one of important reasons which result in proliferation unbalance of cells. 2. p16 gene mutation in exon 2 presents in HCC, but it does not frequently occur in Chinese hepatocarcinogenesis. 3. p16 gene abnormality and cyclinD1 over expression may coact in HCC.

[Relationship between vascular endothelial growth factor expression and microvessel density in hepatocellular carcinomas and their surrounding liver tissue].

The relationship between vascular endothelial growth factor (VEGF) expression and microvessel density was studied with immunohistochemical method in hepatocellular carcinoma (HCC) and pericarcinomatous liver tissue. The positive rate of VEGF in HCCs was significantly lower than in surrounding liver tissues (66.7% vs. 85.4%, P < 0.05). There was no significant difference between HCC and pericarcinomatous liver tissue on expressive intensity of VEGF. The positive signal of VEGF was mainly localized in cytoplasma of cancer cells, pericarcinomatous hepatocytes, and vascular endothelial cells. The microvessel density in HCC was higher than in pericarcinomatous liver tissue and closely correlated to differentiated degree of cancer cells (rs = 0.5870; rs = 0.8235). The poorer cancer cell differentiation, the higher microvessel density. The results suggest that VEGF may not be the sole factor that stimulates angiogenesis in HCC genesis and development. To detect microvessel density in judging prognosis and biological behavior of HCC is more important than that of VEGF.

[Effect of activation of p-MAPK on activating c-fos and c-jun proteins in breast cancer].

OBJECTIVE: This paper was to investigate the relationship between expression of p-MAPK and oncogenesis of breast cancer. METHODS: Immunohistochemical technique was used to detect the expression of p-MAPK and c-fos and c-jun proteins in 68 cases of breast cancers, 42 cases of pericarcinomatous tissues and 7 cases of normal breast tissues. RESULTS: Positive stainings of p-MAPK, c-fos, and c-jun were localized in cancer cell nuclei. The positive rates of p-MAPK, c-fos, and c-jun were 86.8% (59/68), 82.4% (56/68), and 77.9% (53/68), respectively, which were much higher than that in pericarcinomatous tissues (P < 0.01). Of some cases, p-MAPK positive staining was also found in the nuclei of the fibroblastic and angioendothelial cells of cancer stroma. CONCLUSION: Activated or overexpressive MAPK activates the immediately-early oncogenes(c-fos, c-jun), which might play an important role in carcinogenesis of breast cancer and be an early event of oncogenesis of breast cancer.

Microtubule and microfilament distribution and tubulin content in the cell cycle of Indian muntjac cells.

Using DAPI, rabbit antitubulin antibody, FITC-labeled goat anti-rabbit IgG, and TRITC-phalloidin to stain individual cells, the microspectrophotometric analysis showed that three markers that represent the nucleus, microtubules (MT), and microfilaments (MF), respectively, could be recognized in individual cells without interference. The phase of the cell cycle was determined by DNA content. We found that in Indian muntjac (IM) cells, the amount of tubulin in G2 and M phases was about twice as much as that in G1 phase. In G2 cells, the cytoplasmic microtubule complex (CMTC) became denser than in G1 cells. The cytoplasmic MT extent in basically the same orientation as MF bundles in interphase. The regions where the MT is denser also have a denser MF distribution.