[Advances in the molecular genetic studies of acephalic spermatozoa syndrome].

Acephalic spermatozoa syndrome (ASS) is characterized by a predominance of headless spermatozoa with abnormal head-tail junction in the ejaculate, which causes severe male infertility. The pathogenic mechanism of ASS remained unclarified for a long time until recent identification of the four ASS-associated genes SUN5, PMFBP1, TSGA10, and BRDT and their mutations due to the development of high-throughput sequencing technology. This review summarizes the advances in the genetic studies of ASS, focusing on its pathogenic molecular mechanisms, which provide an important basis for the molecular diagnosis of the disease as well as for assisted reproductive technology.

[Expression and diagnosis value of Fascin in non-small cell lung cancer patients].

OBJECTIVE: To evaluate the diagnostic significance of Fascin protein in non-small cell lung cancer (NSCLC) patients. METHODS: Among 110 cases of NSCLC patients and 50 cases of healthy controls, double antibody sandwich enzyme-linked immunosorbent assay (ELISA) was employed to measure the serum levels of Fascin protein, carcinoembryonic antigen (CEA), neuron-specific enolase (NSE), scales like squamous cell carcinoma (SCC) antigen, cytokeratin protein 21-1 (CYFRA21-1) and gastrin-releasing peptide (GRP) precursor. Fascin-positive rate (24.4%) in NSCLC patients was significantly higher than that in healthy controls (4%) (P < 0.05).In different clinical stages of non-small cell lung cancer patients, Fascin-positive rate was statistically significant (P < 0.05). The positive rates of Fascin for CEA, CYFRA21-1, NSE, SCC and Pro-GRP were 25.64%, 5.1%, 5.1%, 5.0% and 1.7% respectively. CONCLUSION: As a new tumor marker, Fascin may be used for clinical screening and prognosis in lung cancer.

Expression and clinical significance of mTOR in surgically resected non-small cell lung cancer tissues: a case control study.

AIMS: Mammalian target of rapamycin (mTOR) is master regulator of the PI3K/Akt/mTOR pathway and plays an important role in NSCLCs. Here we characterized mRNA and protein expression levels of mTOR and its functional associated molecules including PTEN, IGF-1R and 4EBP1 in surgically resected NSCLCs. METHODS: Fifty-four patients with NSCLCs who underwent pulmonary resection were included in current study. mRNA levels of mTOR, PTEN, IGF-1R, and 4EBP1 were evaluated by RT-PCR and protein expression of mTOR, PTEN, and IGF-1R by immunohistochemistry (IHC). Association of expression of the relevant molecules with clinical characteristics, as well as correlations between mTOR and PTEN, 4EBP1 and IGF-1R were also assessed. RESULTS: The results of RT-PCR showed that in NSCLCs, the expression level of mTOR increased, while PTEN, 4EBP1 and IGF-1R decreased. Statistical analysis indicated high IGF-1R expression was correlated with advanced clinical stage (stage III) and PTEN expression was reversely associated with tumor size (P=0.16). The results of IHC showed mTOR positive staining in 51.8% of cases, while IGF-1R positive staining was found in 83.3% and loss of PTEN in 46.3%. Protein expression of mTOR was correlated with its regulators, PTEN and IGF-1R, to some extent. CONCLUSIONS: Abnormal activation of mTOR signaling, high expression of IGF-1R, and loss of PTEN were observed in resected NSCLC specimens. The poor expression agreement of mTOR with its regulators, PTEN, and IGF-1R, implied that combination strategy of mTOR inhibitors with other targets hold significant potential for NSCLC treatment.

[Relationship between vascular endothelial growth factor C expression level and lymph node metastasis in non small cell lung cancer].

OBJECTIVE: To investigate the relationship between vascular endothelial growth factor C (VEGF-C) expression level and lymph node metastasis in non small cell lung cancer (NSCLC). METHODS: Fifty-two NSCLC patients, 38 males and 14 females, aged (59+/-11) (29-77), were divided into 2 groups based on the pathological examination: lymph node metastasis positive group (n=25) and lymph node metastasis negative group (n=27). RT-PCR and immunohistochemistry were used to detect the mRNA and protein expression of VEGF-C in the tumor tissues and lymph nodes resected during operation. RESULTS: The VEGF-C mRNA expression level in the lung tumor tissue of the lymph node metastasis positive group was 0.273+/-0.179, significantly higher than that in the lymph node metastasis negative group (0.089+/-0.087, P<0.01). The VEGF-C mRNA expression level in the positive lymph node of the lymph node metastasis positive group was 0.207+/-0.174, significantly higher than that in the lymph node metastasis negative group (0.114+/-0.107, P<0.01), but not significantly different from that in the negative lymph nodes of the same group(0.196+/-0.186, P>0.05). The VEGF-C protein positive rate of the lung cancer tissues of the lymph node metastasis positive group was 93.3% (14/15), significantly higher than that of the lymph node metastasis negative group (6.7%, 1/15, P<0.01). The VEGF-C protein positive rate of the metastasis positive lymph noses was 80.4% (37/46), and all 52 metastasis negative lymph nodes were VEGF-C protein negative. CONCLUSION: VEGF-C mRNA and protein expression levels predict lymph node metastasis and can be useful predictors of lymph node metastasis in NSCLC.

[Ectogenous fragile histidine triad gene inhibits the malignant phenotype of human lung cancer cell line A549 in vitro and in vivo].

OBJECTIVE: To investigate the inhibition effects of fragile histidine triad (FHIT) gene on the malignant growth of A549 cell line. METHODS: A mammalian expression vector PEGFP-FHIT was constructed and transfected into the A549 cell line by lipofectamine. Then the transfected cell line was screened by G418. Individual G418-resistant colonies were isolated by limited dilution. The monoclonal transfected cell line was screened by RT-PCR and immunochemical staining. The inhibition growth efficacy of extraneous FHIT was evaluated by clonogenic survival assay, flow cytometry and heteroplastic transplant on nude mice. RESULTS: Presence of extraneous FHIT gene in FHIT-A549 cell was proved by RT-PCR. Immunochemical stain demonstrated that the expression of extraneous FHIT protein was positive in FHIT-A549 cell and negative in PEGFP-A549 cell and A549 cell. The clonal formation rate of FHIT-A549 (2.6%) was significantly lower than that of A549 cell (50.1%) and PEGFP-A549 cell (53.6%, P < 0.01). FHIT-A549 cell (95.8%) was blocked in G(2) phage. Tumorigenicity of A549 cells in nude mice was greatly inhibited by expression of ectogenous FHIT gene. The weight of tumor was significantly lower in FHIT-A549 cell (0.04 +/- 0.03) than in A549 cell (0.24 +/- 0.11) and PEGFP-A549 cell (0.25 +/- 0.07, P < 0.01). CONCLUSIONS: Reintroduction of the expression of ectogeneous FHIT gene can obviously suppress the proliferation and tumorigenicity in human lung cancer cell line A549 and induce apoptosis. The data demonstrate oncosuppressive properties of FHIT gene.

[Growth inhibitory effect of adriamycin conjugated to single-chain antibody on human lung adenocarcinoma in vitro].

OBJECTIVE: To evaluate the growth inhibitory effect of adriamycin (ADM) conjugated to an anti-lung cancer single-chain antibody (ScFv) 2A7-1 on lung adenocarcinoma cell line A2 in vitro. METHODS: 2A7-1 cell culture medium was concentrated by ultra-filtration (with Amicon P10Z filter), and soluble ScFv was purified using RPAS purification kit. ADM was conjugated to 2A7-1 by glutaraldehyde. A(280) and A(490) of the conjugate 2A7-1-ADM were determined by spectrophotometry and the molar ratio of 2A7-1 to ADM was calculated. Immunoreactivity of the conjugate was detected by immunohistochemistry. Its growth inhibitory effect on lung adenocarcinoma cell line A2 was determined by colony formation assay in vitro. RESULTS: The molar ratio of 2A7-1 to ADM was 1:3.2. The conjugate strongly reacted with A2 cell. Its growth inhibitory effect on A2 cells was 4 times as potent as ADM. CONCLUSION: Adriamycin conjugated to anti-lung cancer single-chain antibody 2A7-1 has much higher cytotoxic activity than unconjugated adriamycin against human lung adenocarcinoma.

[Inhibitory effect of p53 with deletion of c-terminal 356 - 393 amino acids on malignant phenotype of human lung cancer cell line].

OBJECTIVE: To study the effect of extraneous p53 gene with deletion of c-terminal 356 - 393 amino acids on inhibition of malignant phenotype of human lung cancer cell line. METHODS: Recombinant plasmid pEGFP-p53 (del) with codon deletion of c-terminal 37 amino acids from 393 to 356 region and pEGFP-p53 (wild type) were constructed. The human lung cancer cell line 801D served as a receipt cell had p53 deletion and mutation at 248 codon. 801D cells, having been transfected by pEGFP-p53 (wild type), pEGFP-p53 (del) or pEGFP, were selected by G418. Growing transfected cells were cloned respectively by method of dilution. Presence of extraneous gene was detected by PCR, their expression in cells was examined by fluorescence microscopy. Cloning efficiency was in vitro tested to examine the cellular proliferating ability. The xenograft in nude mice was performed and xenograft tumors were weighed one month later. Expression of GFP in tumor and transplanted cellular mass were detected by blot slices. RESULTS: pEGFP-p53 (del)-801D, pEGFP-p53-801D and pEGFP-801D were established. Extraneous p53 gene and expression of GFP were found in pEGFP-p53 (del)-801D and pEGFP-p53-801D. Inhibitory rate of colony was 99.6% for pEGFP-p53 (del)-801D and 81.0% for pEGFP-p53-801D. Inhibition of malignant proliferation of extraneous p53 (del) was higher than that of p53 (wild type) (P < 0.01). Even when inhibition of malignant proliferation extraneous pEGFP-p53 (del) was obvious, 0.2% colonies were formed, extraneous p53 and expression of GFP were observed. Animal test showed that tumor on the nude mice was positive (4/4, 4/4) in the control group (801D and pEGFP-801D), but negative (0/4, 0/4) in the experiment group [pEGFP-p53 (del) 801D and pEGFP-p53 (wild type) 801D]. Expression of GFP in the cells of cellular mass transplanted by pEGFP-p53 (del) 801D or pEGFP-p53 (wild type) 801D was observed. CONCLUSION: In vitro inhibitory effect of extraneous p53 gene with deletion of C-terminal 356 - 393 amino acids on malignant growth of lung cancer cell with p53 mutation or deletion at 248 codon is marked. Inhibitory action of p53 on malignant proliferation of cancer cells is heterogeneous.