临床肿瘤细胞学研究的历史回顾与展望

中国临床肿瘤细胞学(Chinaclinicaloncocytology)工作起始于20世纪50年代初,历经了半个多世纪艰难曲折的发展。在各种新技术、新方法的推动下出现了突飞猛进的局面。巴氏涂片的临床应用也始于当时。由于全国开展了宫颈癌的普查,宫颈癌的发病率和死亡率都有明显的下降。在各种宫颈癌筛查试验中,液基细胞学检查的诊断价值最高。细针穿刺细胞学的应用,目前已遍及人体各个脏器和部位,根据数万例的观察,尚未发现肿瘤沿针道扩散的证据。过去30年间,我国进行食管拉网细胞学检查36万人次,检出食管上皮细胞重度增生近万例,食管癌和贲门癌27000余例,早期癌2800多例,食管拉网法发现无症状的早期食管癌占食管癌的80%以上,食管拉网法的阳性准确率高达98.5%。核仁组成区嗜银染色的临床应用十分广泛,我国发表的有关这方面的研究论文已超过500篇,内容包括常见的各种肿瘤,如:胃、肺、肝、大肠、乳腺等各种恶性肿瘤,在良恶性肿瘤的鉴别诊断中以恶性淋巴瘤和淋巴结良性疾患的鉴别最有价值。对临床肿瘤细胞学工作的今后发展,本文提出了四点建议。

Relationship between proliferative activity of cancer cells and clinicopathological factors in patients with esophageal squamous cell carcinoma

AIM: To assess whether the molecular markers of malignant tumors could improve the understanding of tumor characteristics, and to observe the characteristics of expression of cell cycle markers Ki-67 and cyclin A in esophageal carcinoma and to analyze the relationship between proliferative activity of cancer cells and clinicopathological factors. METHODS: Seventy of surgically resected esophageal squamous cell carcinoma (SCC) were examined by immun-ohistochemistry utilizing commercially available antibodies. Nuclear staining was regarded as a positive result. At least 50 fields in each tumor and non-tumor section were evaluated at a medium power (×200) to determine the proportion of tumor cells and the staining intensity of nuclei in the entire sections. RESULTS: Ki-67 and cyclin A were only expressed in base cells of normal esophageal mucosa. The positive immuno-staining of nuclei of SCC was significantly higher than that in normal esophageal mucosa (t = 13.32 and t = 7.52, respectively, P<0.01). The distribution of positively stained was more diffuse and stronger in poorly differentiated SCC. Both Ki-67 and cyclin A expressions were related to histological grades of tumors (t = 3.5675 and t = 3.916; t = 2.13, respectively, P<0.05) but not to the sex and age of the patients, tumor size, lymphatic invasion, location, or stage grouping. CONCLUSION: The proliferative activity of cancer cells may be understood by immunohistochemistry of Ki-67 and cyclin A in Chinese patients with esophageal SCC. These cell cycle markers may serve as an indicator of cancer cell proliferation rate. The overexpression of cell cycle markers Ki-67 and cyclin A suggests the poor SCC differentiation in patients with esophageal carcinoma.

Cytotoxic T-lymphocytes response in vitro activated by dendritic cells pulsed with heat shock protein 70 derived from human bladder tumor cell lines of EJ

Objective： To investigate whether human dendritic cells （DC） derived from peripheral blood mononuclear cells （PBMC）, which were pulsed by heat shock protein 70 （HSP70） isolated from human bladder tumor cell lines of E J, were able to induce peptide specific cytotoxic T-lymphocytes （CTL） response in vitro and give the experimental foundation for the future clinical trials of immunotherapy in bladder tumor. Methods： The E J-derived HSP70 co-cultured with DC from the healthy volunteers＇ PBMC, along with the crude lysate （the supematant before HSP70 purification） from EJ cells were used as the experimental groups and DC not pulsed by any tumor cells antigen were the blank control. The autologous T-lymphocytes were added into the above various DC groups, and after incubation, the stimulation indexes （SI） and interferon-y （IFN-γ） were detected to evaluate the immune activities of various DC groups. The killing effects of CTL to target cells, EJ and Hela cells, were determined with 51^Cr releasing test. Results： Both DC/HSP70 and DC/the crude lysate could effectively activate CTL in vitro and kill target cells EJ. The killing effect of DC/HSP70 to EJ was much stronger than DC/the crude lysate （the supernatant before HSP70 purification） （P 〈 0.05）. DC without any tumor cell antigens had a lower killing power to EJ. Meanwhile, DC/ HSP70 had little killing power to Hela non-relevant to bladder tumor histopathologically as compared with EJ cells （P 〈 0.05）. Conclusion： The DC pulsed by HSP70 derived from the autologous tumor cells could induce a peptide complexes specific CTL response to tumor cells, and the CTL response induced by the DC/HSP70 was stronger, which display the basis of the possible clinical application of DC/HSP70 for bladder tumor.

Construction of eukaryotic expression plasmid pEGFP-N1-WWOX and its transient expression in SMMC-7721 cells

Objective： To construct eukaryotic expression plasmid pEGFP-NI-WWOX and transiently express it in SMMC-7721 cells. Methods： Total mRNA was extracted from normal human liver tissue. RT-PCR was used to amplify the aimed segments WWOX cDNA which was then digested with Hindlll and BamHI and inserted into a eukaryotic expression plasmid pEGFP-N 1 to construct pEGFP-N 1-WMVOX. The constructed plasmid was transfected into SMMC-7721 cells by lipofectamine 2000 - mediated transfer method. The expression of WWOX in transfected SMMC-7721 cells was detected 24, 36 and 48 h post-transfection with fluorescence microscope and the expression level of WWOX mRNA in transfected SMMC-7721 cells was assay by using RT-PCR. The change of MMWOX expression and cell proliferation rates were detected by immunocyto- chemistry and MTT methods respectively. Results： The results showed pEGFP-N1-WWOX was successfully constructed and expressed transiently in SMMC-7721 cells. At 48th hour post-transfection, the number of positive cells was increased significantly and much brighter green fluorescence could be detected, while no green fluorescence was detected in the control group. In SMMC-7721 cells transfected with pEGFP-NI-WWOX a high level of porcine WWOX was detected. WWOX ex- pressed by transfected cells could significantly inhibit the proliferation of SMMC-7721 cells. Conclusion： pEGFP-N1-WWOX was expressed successfully in SMMC-7721 cells, which suggested that might be used as a new therapeutic method for liver cancer.

Overexpression of β3/γ2 chains of laminin-5 and MMP7 in biliary cancer

AIM： To clarify the clinicopathological significance of laminin-5 y2 （LNγ2） and 133 （LNI33） chains and MMP7 expression in biliary tract cancer. METHODS： We analyzed the association between immunohistochemically detected LNγ2, LNβ3, and MMP7 expression in biliary tract cancer and clinicopathological characteristics. Activity of MMP7 was analyzed by casein zymography. An in vitro invasion assay after treatment with MMP7-specific siRNA was performed. RESULTS： LNγ2 expression was predominantly observed in carcinoma cells at the invasive front. LNγ2 expression was seen in 57% of patients with biliary tract cancer, and was associated with depth of invasion, histologic type, and advanced stage. The expression pattern of LNβ3 was classified Into two types： invasive front dominant type （38%） and diffuse type （28%）.The invasive front dominant type was associated with histologic type and advanced stage. MMP7 positivity was correlated with LNγ2 or LNβ3 expression but not with clinicopathological characteristics. Active MMP7 detected by casein zymography was correlated with depth of invasion and advanced stage. Downregulation of MMP7 expression by siRNA resulted in a significant decrease in biliary tract cancer cell invasion in vitro.CONCLUSION： Our results suggest that LNγ2 and LNβ3, in conjunction with MMP7, play a key role in the progression of biliary tract cancer.

Research of cervical cytology screening in diagnosis of 370,000 cases with cervical lesion in Guangdong

Objective： Analysis of the results of 37 million cases by using cervical liquid-based cytology screening, discussion the situation of cervical cancer and precancerous lesions in Guangdong province. Methods： From 2002 to 2006, using liquid-based thin-layer cytology （TCT） method screening cervical lesions within 371,929 women in Guangdong. Cytological diagnosis adopt TBS （the Bethesda system） Standard （TBS improved 2001 version of the diagnostic criteria）, statistic positive rate by age and region respectively. Results： 371,929 cases of gynecology cervical samples in Guangdong, 331,251 cases were negative, accounted for 91.23%; cytology positive cases： 32,548 cases were squamous intraepithelial lesions （8.96%）, 486 cases were glandular epithelial lesions （0.13%）; grouped according to age： 30 to 40-year-old positive rate was 9.13%, 40 to 50-year-old positive rate was 9.60%, the latter had higher cytology positive rate; grouped according to region： the total samples in PRD areas were 304.951 cases, accounted for 81.99%, in which 24301 cases were positive （8.14%）, in 66978 cases （18.01%） of NON-PRD regions, 7645 cases were positive （11.87%）, there were 1858 cases had biopsy/follow-up results from 2004 to 2005, and the rate was 2.88%. Conclusion： The cytological positive rate of cervical cancer and its precancerous lesions was 9.09% in Guangdong （of which the rate of cytological diagnosis as precancerous lesions was 3.40%）, 30 to 50 age group had the high incidence of cervical cancer and its precursor lesions, and also the main stage of prevention for cervical cancer. Cytology positive rate was 11.87% in economic underdevelopment Non-Pearl River Delta regions, much higher than the Pearl River Delta region.

Properties and feasibility of using cancer stem cells in clinical cancer treatment

Cancer treatment failure, drug resistance, or metastatic recurrence are thought to be caused mainly by the existence of a very small number of cancer stem cells(CSCs). The characteristics of this subgroup of cells include self-renewal, tumorigenesis, multiple differentiation and high invasiveness, metastasis, and drug resistance potential. Many studies have demonstrated that CSCs play important roles in tumor growth, spread and metastatic relapse after treatment, and are closely related to the prognosis of patients.From a therapeutic viewpoint, deep insights into the CSCs biology, development of specific therapeutic strategies for targeting CSCs, and characterization of their microenvironment could be an ideal way to combat cancer.

The diagnostic evaluation of fine needle aspiration cytology of thyroid and its clinical application

Objective:The aim of the study was to investigate the diagnostic value of fine needle aspiration cytology (FNAC) and its clinical application.Methods:From April 2009 to February 2011,thyroid FNAC were performed in a total of 186 patients with thyroid nodule or mass in our hospital and 78 of those 186 patients subsequently underwent thyroidectomy.The FNAC findings were compared with the results of the corresponding histological diagnosis.Results:The results of thyroid FNAC for 186 patients showed that,(1) 166 cases of benign lesions,the detection rate was 89.24% (166/186),including 96 cases of nodular colloid goiter (51.61%),28 cases of simple colloid goiter (15.05%),38 cases of Hashimoto's thyroiditis (HT) (20.43%) and 4 cases of thyroid adenoma (2.15%);(2) 4 cases of suspicious malignant lesion,the detection rate was 2.15% (4/186);(3) 16 cases of malignant tumor,the detection rate was 8.60% (16/186).Seventy eight patients including malignant (16),suspicious malignant (4),HT (20) and nodular colloid goiters (38) cases diagnosed by FNAC were performed operation with thyroidectomy and the postoperative histopathologic results showed that there were 2 cases HT combined thyroid papillary carcinoma in HT 20 cases by FNAC,15 cases of thyroid papillary carcinoma and 1 case of follicular carcinoma in 16 cases of malignant tumor by FNAC and 4 case of thyroid papillary carcinoma in 4 cases of suspicious malignant by FNAC.Conclusion:Thyroid FNAC is a valuable and reliable method for the diagnosis of the thyroid nodules or mass or even most diffuse thyroid diseases.Diagnosis of HT and thyroid papillary carcinoma can be made by thyroid FNAC.There was larger hint value for nodular colloid goiter and simple colloid goiter according to thyroid FNAC.

Role of integrin oV in sensitivity to radiotherapy of human NPC

Objective： To investigate whether the cell adhesion molecule integrin αV could influence the radiosensitivity of human nasopharyngeal carcinoma CNE-2Z cells via effecting intercellular effect. Methods： Human NPC cell line CNE-2Z was cultivated as three dimensional model using liquid overlay technique. Cell counting was employed to detect the radiosensitivity of CNE-2Z cells by blood cell counter before and after blocking integrin aV function; cell apoptosis was detected by flow cytometry using Annexin V/PI label. Results： Three dimensional culture model of human NPC cell line CNE-2Z cells were successfully established. The integrin αV expression of CNE-2Z multicellular spheroids elevated as the dosage of radiotherapy increased. Blocking of integrin aV function leaded to higher radiosensitivity and more apoptotic cells of CNE-2Z multicellular spheroids. Conclusion： Cell adhesion molecules integrin aV can influence the radiosensitivity of human nasopharyngeal carcinoma CNE-2Z cells and the inhibition of cell apoptosis might be its mechanism, proved by three dimensional culture model to mimic solid tumor in vivo.

Clinicopathological Analysis of a Primary and Solitary Brain Plasmacytoma: Case Report with an Encepha-locoele

A primary and solitary plasmacytoma with an encephalocoele is an extremely rare tumortx. The origin of plasmacytoma cells has not been clarified, but a number of studies have suggested the possibility that it originates from the meninges and reticular cells of the Virchow-Robin space. Other studies have suggested that plasmacytoma cells may possibly arise from original reticular and stem cells.