Human dermal fibroblast isolation using two enzyme digestion methods: a comparative study / 中国组织工程研究

BACKGROUND: Dermal fibroblasts are widely used and demanded, and there are various isolation methods, but no comparative studies on enzyme digestion methods are reported. OBJECTIVE: To compare the cell count, morphology, migration and proliferation of human dermal fibroblasts isolated by two enzyme digestion methods. METHODS: Human dermal fibroblasts were isolated using either dispase-collagenase or trypsin, and their cell yield and viability were assessed by morphology, cell count and proliferation curve by cell counting-kit 8 assay. The ability of migration was observed by cell scratch test. RESULTS AND CONCLUSION: The fibroblasts digested with dispase-collagenase were fused at 6-7 days after inoculation, and the cells isolated by trypsin digestion were fused at 8-9 days after inoculation. Fibroblasts could be obtained by both two digestion methods. The production in the dispase-collagenase group was significantly higher than that in the trypsin group. The migration rate in the dispase-collagenase group was significantly faster than that in the trypsin group. The growth cures of the human dermal fibroblasts in the two groups revealed that the cell count was positively correlated with time, and the absorbance values of the dermal fibroblasts in the dispase-collagenase group were significantly higher than those in the trypsin group at 3, 4 and 5 days after incubation. To conclude, the cell yields, migration and proliferation of dermal fibroblasts digested with dispase-collagenase are significantly higher than those of the cells digested by trypsin, indicating that dispase-collagenase digestion results in better isolation and viability of dermal fibroblasts from the dermis.

Morphology, immunohistochemistry and hTERC gene in-situ hybridization in Barrett's esophagus / 中华病理学杂志

<p><b>OBJECTIVE</b>To study the clinicopathologic features and differential diagnosis of proximal gastric mucosa and mucosa of Barrett's esophagus (BE) in biopsy specimens.</p><p><b>METHOD</b>Thirty-eight cases of Barrett's esophagus (diagnosed using WHO criteria) and 44 cases of proximal gastric mucosa were studied by immunohistochemistry (for CK7, CK20, CK4, CK8, S-100 protein, MUC6, COX2 and bcl-2) and fluorescence in-situ hybridization (FISH) (for hTERC gene). The pathologic features were analyzed.</p><p><b>RESULTS</b>The differences in expression of CK7, CK20, MUC6, COX2 and bcl-2 between BE and proximal gastric mucosa with intestinal metaplasia were not statistically significant (P > 0.05). There was however a statistically significant difference in expression of S-100 protein (P < 0.05). The expression of CK7/CK4 and CK7/CK8 in BE showed positive correlation (P < 0.05). However, such correlation was not demonstrated in proximal gastric mucosa (P > 0.05). The results of hTERC gene expression by FISH showed a statistically significant difference between the two groups: 57.9% (22/38) in BE and 13.6% (6/44) in proximal gastric mucosa (P < 0.05).</p><p><b>CONCLUSIONS</b>The significance of CK7 and CK20 expression is uncertain in the differential diagnosis between BE and proximal gastric mucosa. On the other hand, positivity for CK7/CK4/CK8 may support the diagnosis of BE and play a role in distinguishing between the two groups. S-100 protein expression and detection of hTERC gene amplification also contribute to the diagnosis of BE.</p>

Serrated lesions of colon and their malignant potential / 中华病理学杂志

<p><b>OBJECTIVE</b>To study the serrated lesions of colon and to compare the malignant potential between traditional serrated adenomas (TSA) and conventional adenomas (CAD).</p><p><b>METHODS</b>A total of 5347 cases of colorectal polyps encountered in five regional hospitals during a five-year period were retrospectively reviewed. The serrated lesions were classified on the basis of histologic examination. One hundred and eighty-seven cases of CAD (including 160 cases of tubular adenoma and 27 cases of villous adenoma) and 36 cases of invasive adenocarcinoma were randomly selected as the controls. The degree of dysplasia and expressions of Ki-67, p53 and beta-catenin in TSA and CAD were compared.</p><p><b>RESULTS</b>Amongst the 5347 colorectal polyps studied, 258 cases (4.8%) of serrated lesions were found, which included 112 cases (43.4%, 112/258) of hyperplastic polyp, 78 cases (30.2%, 78/258) of TSA and 26 cases (10.1%, 26/258) of sessile serrated adenoma. Sixty-two cases of TSA were identified from 3 hospitals, in which moderate dysplasia was found in 13 cases. High-grade intraepithelial neoplasia and ICA were found in 6 cases (9.6%). Compared with the 187 cases of CAD, moderate dysplasia were found in 27 cases and high-grade intraepithelial neoplasia and invasive adenocarcinoma were found in 25 cases (13.3%, χ(2) = 19.373, P = 0.000). There was statistically significant difference between TSA and CAD in the degree of dysphasia. The expression of Ki-67, p53 and beta-catenin in TSA and CAD showed no significant difference (P > 0.05).</p><p><b>CONCLUSIONS</b>The incidence of serrated lesions is lower in northern Chinese population than that in Caucasians. TSA has obvious malignant potential; but the rate associated with high-grade intraepithelial neoplasia and invasive adenocarcinoma is lower than that in CAD.</p>

Clinicopathologic features and proliferative status of colorectal serrated lesions: a study of 104 cases / 中华病理学杂志

<p><b>OBJECTIVE</b>To study the clinicopathologic features and proliferative status of colorectal hyperplastic polyp (HP), sessile serrated adenoma (SSA) and traditional serrated adenoma (TSA).</p><p><b>METHODS</b>One hundred and four cases colorectal serrated lesions were collected from 2628 cases of colorectal polyps during the period from November, 2002 to December, 2007. The clinicopathologic features and expression of proliferation marker Ki-67 were studied.</p><p><b>RESULTS</b>On the basis of morphologic examination, 60 cases were classified as HP, 20 cases as TSA, 11 cases as SSA, 7 cases as mixed HP/SSA/TSA, and 6 cases as mixed serrated polyp/adenoma and tubular adenoma. Immunohistochemical study for Ki-67 showed that 40 cases (78%) of the 51 cases of HP were either mostly negative or rarely (<25% cells) positive. Most of the positive cells were located at crypt bases. Among the 15 cases of TSA, 11 of them revealed positive cryptal cells (25% to 50% or>50% positivity). Most of the positive cells were located in mid portion of crypts. The number and distribution of Ki-67 positive cells in SSA were similar to those in TSA but were significantly different from those in tubular adenoma and adenocarcinoma (chi2=34.601, P=0.000; chi2=63.077, P=0.000, respectively).</p><p><b>CONCLUSIONS</b>HP, SSA and TSA have their morphologic characteristics, with some overlapping features noted. The distinction between SSA and HP can be difficult. Diagnosis of SSA relies mostly on architectural rather than cytologic features. The distinction between TSA and SSA depends mainly on the presence of dysplasia. Ectopic crypt formation is almost exclusively seen in TSA. The distribution and percentage of Ki-67-positive cells are also helpful in subtyping of various colorectal serrated lesions. In general, the proliferative index is lower in serrated adenoma (TSA or SSA) than in tubular adenoma.</p>

Clinicopathological and immunohistochemical features of colorectal sessile serrated adenoma / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To investigate the clinicopathological characteristics and expression status of Ki67, p53, CEA, CDX, CK7 in colorectal sessile serrated adenoma (SSA).</p><p><b>METHODS</b>The clinicopathological data of 11 cases of SSA, 51 cases of hyperplastic polyp (HP) and one case with mixed HP/SSA were reviewed and analyzed retrospectively. The expression of Ki67, p53, CEA, CDX and CK7 were detected by immunohistochemistry.</p><p><b>RESULTS</b>The major histological features in SSA were architectural abnormality in crypts, dilatation of serrated crypt bases like an inverted "T" or "L" shape adjacent to muscularis mucosa. Atypical cells containing round to oval nuclei and nucleoli were also observed. The immunohistochemical staining showed that the expression of p53 increased gradually from HP to TA: 11.8% in HP, 20.0% in SSA, 41.2% in VTA and 75.0% in TA, with a significant difference among the groups (chi(2) = 17.996, P = 0.000). However, no significant difference in the expression of CDX and CK7 was observed between HP and SSA. Of the 10 SSA cases, positive expression of Ki67 was found in cells located in the base or middle part of crypt in 6 cases, positive cells index was 26% - 50% in 5 cases, and > 50% in 3. Compared with the expression of Ki67 in the HP, VTA and VA, SSA showed a significant difference in both the positive cell number and in the positive regions. (positive number: chi(2) = 34.601, P = 0.000; positive regions: chi(2) = 63.077, P = 0.000).</p><p><b>CONCLUSION</b>Morphological diagnosis of SSA was mainly based on crypt architectural and cellular abnormalities, and the crypt architectural abnormality may be more important than cellular features. Detection of p53 and Ki67 expression may be helpful in differential diagnosis and understanding the nature of SSA.</p>

Survey on reproductive health status of Shanghai college students / 中华男科学杂志

<p><b>OBJECTIVE</b>To explore reproductive health condition of college students in Shanghai and provide scientific basis for reproductive health education.</p><p><b>METHODS</b>From July 2004 to March 2005, 1192 college students (majors including medicine, science and engineering, arts and culture; 618 medical students and 549 non-medical; 555 male and 612 female) from seven time-day colleges in Shanghai were investigated with a reproductive health questionnaire involving sex. The questionnaire was constituted by 72 topics, involving the need of education, sex attitude, sex videos, sex hygiene knowledge and sexually transmitted diseases (STDs), etc.</p><p><b>RESULTS</b>39.33% (459/1167) students thought they had accepted formal sex education; 49.19% (574/1167) students chose lecture as the favorite way; 41.90% (489/1167) students thought the reproductive health education would not increase the incidence of premarital sexual behavior; 64.10% (748/1167) students thought it was necessary to give college students reproductive health instructions and knowledge. On the other hand, we found 45.05% (250/555) boys and 27.45% (168/612) girls could accept the premarital sexual behaviors; 35.32% (196/555) boys and 39.38% (241/612) girls could accept premarital pregnancy; 44.39% (518/1167) students could calculate the safe period accurately; 71.38% (833/1167) students knew emergency contraception measures; 57.67% (673/1167) students knew how to and when to use condoms; To our surprise, only 2.19% (12/549) non-medical students and 12.14% (75/618) medical students knew exactly the whole 4 sexually transmitted diseases that needed to be essentially prevented in China.</p><p><b>CONCLUSION</b>At present, the reproductive health condition of college students in Shanghai is sophisticated. Much more education on sex safety, self-protection skills and acquired immunodeficiency syndrome (AIDS) knowledge, etc should be reinforced.</p>

Expression of PH20 in primary and metastatic breast cancer and its pathological significance / 中华病理学杂志

<p><b>OBJECTIVE</b>To evaluate the expression pattern of PH20 in primary and metastatic breast cancer and its relationship to tumor metastatic potential.</p><p><b>METHODS</b>Anti-PH20 antibody was synthesized by injection of conjugated human PH20 peptides into rabbits. Immunohistochemical study was performed on 53 cases of human breast cancer. Western blot was used to detect PH20 expression in 5 cases of breast cancer with available fresh tissue. Two oligonucleotide probes were prepared for in-situ hybridization using breast tissue microarray.</p><p><b>RESULTS</b>Normal breast tissue did not express PH20 (0/3), while 58.4% (31/53) of breast cancer cases did. The highest expression rate was found in metastatic foci in regional lymph nodes (83.3%), followed by primary breast cancer tissue in cases with lymph node secondaries (70.8%). The breast cancer cases with no any metastasis had an expression rate of 48.2%. The immunohistochemical staining results were further confirmed by Western blotting. In-situ hybridization showed PH20 RNA in 75% of the breast cancer tissue (21/28). Two of the 17 cases of normal breast tissue showed weak expression in some ductolobular units.</p><p><b>CONCLUSIONS</b>The expression of PH20 has a positive correlation with metastatic potential in breast cancer. It is possible that PH20 may play an important role in the invasive growth and metastasis of breast cancer cells, via mechanisms such as digestion of surrounding stromal tissue and release of FGF-2.</p>