Functional analysis of transgenic cry1Ah-1 maize.

Maize is an important food crop in the world, but the yield and quality of maize have been significantly reduced due to the impact of insect pests. In order to address this issue, the cry1Ah gene was subjected to error-prone PCR for mutagenesis, and subsequently, the mutant cry1Ah-1 gene was introduced into maize inbred line GSH9901 callus using the Agrobacterium-mediated method. The T2 generation transformed plants were obtained by subculture, and 9 transgenic positive plants were obtained by molecular detection which was carried out by PCR, qRT-PCR, Bt gold-labeled immunoassay test strips, Western blot and ELISA. It was found that the Cry1Ah-1 gene could be transcribed normally in maize leaves, of which OE1 and OE3 had higher relative expression levels and could successfully express proteins of 71.94 KD size. They were expressed in different tissues at the 6-leaf stage, heading stage and grain-filling stage, and could ensure the protection of maize from corn borer throughout the growth period. The biological activities of OE1 and OE3 were tested indoors and in the field, and the results showed that in indoors, the corn borer that fed on OE1 and OE3 corn leaves had a mortality rate of 100 % after 3 days; in the field, OE1 and OE3 had strong insecticidal activity against corn borer, reaching a high resistance level. In conclusion, the transgenic cry1Ah-1 maize has a strong insecticidal effect on corn borer, and has a good prospect of commercialization.

Endometriosis-associated endometrioid adenocarcinoma of the fallopian tube synchronized with endometrial adenocarcinoma: A case report.

BACKGROUND: Endometriosis is a common gynecological disorder that affects women of reproductive age. It is characterized by a cancer-like invasion of the extra-uterine endometrium and exhibits a strong association with ovarian clear cell cancer and endometrioid cancer. Endometriosis-associated fallopian tube endometrioid adenocarcinoma synchronized with endometrial adenocarcinoma was rarely reported. CASE SUMMARY: A 49-year-old woman was referred to our hospital complaining about abnormal vaginal bleeding for three years following unsatisfactory medication. Intraoperative frozen sections unexpectedly unveiled an endometrioid cancer of the left fallopian tube with superficial invasion surrounded by diffuse endometriosis synchronized with endometrioid endometrial cancer. CONCLUSION: It was difficult to make a differential diagnosis when confronted with incidental findings of fallopian tube cancer lesions synchronized with endometrial cancer. The key differential diagnosis of primary endometriosis-associated endometrioid adenocarcinoma of the fallopian tube from endometrial adenocarcinoma involvement relies on the pathological identification of malignant transformation in fallopian tube endometriosis disease.

Clinical Performance of the Xpert® CT/NG Test for Detection of Chlamydia trachomatis and Neisseria gonorrhoeae: A Multicenter Evaluation in Chinese Urban Hospitals.

Background: We aimed to evaluate the clinical performance of the GeneXpert® (Xpert) CT/NG assay for the detection of Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) using urine and cervical swabs collected from patients in China. Methods: This study was conducted from September 2016 to September 2018 in three Chinese urban hospitals. The results from the Xpert CT/NG test were compared to those from the Roche cobas® 4800 CT/NG test. Discordant results were confirmed by DNA sequence analysis. Results: In this study, 619 first void urine (FVU) specimens and 1,042 cervical swab specimens were included in the final dataset. There were no statistical differences between the results of the two tests for the detection of CT/NG in urine samples (p > 0.05), while a statistical difference was found in cervical swabs (p < 0.05). For CT detection, the sensitivity and specificity of the Xpert test were 100.0% (95%CI = 96.8-99.9) and 98.3% (95%CI = 96.6-99.2) for urine samples and 99.4% (95%CI = 96.5-100.0) and 98.6% (95%CI 97.5-99.2) for cervical swabs, respectively. For NG detection, the sensitivity and specificity of the Xpert test were 99.2% (95%CI = 94.9-100.0) and 100.0% (95%CI = 99.0-100.0) for urine and 100% (95%CI = 92.8-100.0) and 99.7% (95%CI = 99.0-99.9) for cervical swabs, respectively. Conclusion: The Xpert CT/NG test exhibited high sensitivity and specificity in the detection of CT and NG in both urine and cervical samples when compared to the reference results. The 90-min turnaround time for CT and NG detection at the point of care using Xpert may enable patients to receive treatment promptly.

Isolation and detection of label-retaining cells in a nasopharyngeal carcinoma cell line.

BACKGROUND AND OBJECTIVE: Detection of label retaining cells (LRCs) has been a method to confirm existence of stem cells, and bromodeoxyuridine (BrdU) has commonly been used for labeling. In this study, to verify stem cells in nasopharyngeal carcinoma (NPC), LRCs were established and detected in NPC cell line 5-8F. METHODS: The 5-8F cells were cultured with BrdU and inoculated subcutaneously into nude mice. By immunohistochemistry, immunocytochemistry, and immunofluorescence, BrdU was detected in 5-8F cells and xenograft tumors. RESULTS: BrdU was strongly positive in cells on the 2nd and the 7th day after being added BrdU, while negative when cells were cultured without BrdU. However, only sporadic cells were positive on the 14th day after BrdU being washed out, and these cells were thought to be LRCs. The average percentage of LRCs was (0.67 +/- 0.32)%. After being cultivated with BrdU for 48 h, 5-8F cells were inoculated into nude mice subcutaneously. After chasing 8 weeks, only sporadic LRCs were detected in xenograft tumors, with a proportion of (0.55 +/- 0.36)%, and these LRCs were located at cancer margin. CONCLUSION: The existence of LRCs in 5-8F cells indicates the existence of cancer stem cells in NPC.

ZEB1 promotes invasion and metastasis of endometrial cancer by interacting with HDGF and inducing its transcription.

Zinc finger E-box binding homeobox 1 (ZEB1), as a typical transcription inhibitory factor of E-cadherin, plays a major role in stimulating the invasion and metastasis of tumors via modulating the epithelial-mesenchymal transition (EMT) signal. However, its function and modulatory mechanisms in endometrial carcinoma (EC) remain unclear. In this study, silencing ZEB1 significantly reduced EC cell migration, invasion, and metastasis, as well as enhanced the sensitivity of EC cells to cisplatin (cDDP) in vitro and in vivo. Mechanism analysis indicated that ZEB1 interacts with hepatoma-derived growth factor (HDGF) and co-localizes in the nucleus. In addition, ZEB1 as a transcription factor binds to the promoter of HDGF to stimulate HDGF transcription. Furthermore, suppression of HDGF in ZEB1-overexpressed EC cells not only reduced the expression of ß-catenin, TCF4, and ZEB1, but also repressed ß-catenin translocation from the cytoplasm into the nucleus and further downregulated the combination with TCF4. Interestingly, the ß-catenin/TCF4 signaling feedback stimulates ZEB1 transcription and therefore constitutes a positive feedback loop. In clinical samples, ZEB1 positively correlates with HDGF expression, and co-expression of ZEB1 and HDGF promotes the pathogenesis of EC. In summary, our study demonstrated that the positive feedback loop of ZEB1/HDGF/ß-catenin/TCF4 plays an unfavorable role in the metastasis of endometrial carcinoma.

[Expression of phosphoglycerate kinase 1 in endometrial carcinoma and its association with patients' outcome].

OBJECTIVE: To investigate the expression of phosphoglycerate kinase 1 (PGK1) and its prognostic value in endometrial carcinoma (EC). METHODS: The expression of PGK1 was detected immunohistochemically in 30 normal endometrium and 130 EC specimens. The relationship between PGK1 protein expression and the clinicopathological features of the patients was evaluated. RESULTS: Immunohistochemical analysis revealed low PGK1 expression in 55.4% (72/130) and high PGK1 expression in 44.6% (58/130) of the EC specimens, as compared with the rates of 90% (27/30) and 10% (3/30) in normal endometrium, respectively (P<0.001). PGK1 expression was significantly correlated with FIGO stage (P<0.001), histological grade (P=0.002) and lymph node metastasis (P<0.001). Kaplan-Meier survival analysis indicated that patients with a high PGK1 expression had a shorter overall survival rate than those with a low PGK1 expression (P<0.001). Multivariate analysis showed that a high PGK1 expression was not the independent predictor of the prognosis of EC (P=0.077). CONCLUSION: A high expression of PGK1 is associated with aggressive and metastatic behaviors of EC, and detection of PGK1 provides assistance in evaluating the prognosis of patients with EC.

[Coexpression of MAP2K4 and vimentin proteins in human endometrial carcinoma and its clinicopathological significance].

OBJECTIVE: To analyze the expression of MAP2K4 and vimentin in human endometrial carcinoma (EC) and their association with the clinicopathological features and prognosis of the patients. METHODS: MAP2K4 and vimentin expressions were detected immunohistochemically in paraffin-embedded tissue sections from 128 patients with EC, and the correlation of MAP2K4 and vimentin expressions with the clinicopathological factors of the patients was analyzed. RESULTS: MAP2K4 and vimentin proteins were positively expressed in 49 (38.3%) and 83 (64.8%) of the patients, respectively. A positive expression of MAP2K4 was negatively correlated with FIGO stage of the tumor (P=0.010) and lymph node status (P=0.016); a positive expression of vimentin was positively correlated with FIGO stage of the tumor (P=0.025), histological grades (P=0.017), depth of myometrial invasion (P=0.044) and lymph node status (P=0.032). MAP2K4 was inversely associated with vimentin expression in EC(r=-0.598, P<0.001). Patients positive for MAP2K4 tended to have a higher overall survival rate (P=0.002), and those positive for vimentin tended to have a lower overall survival rate (P=0.007); patients positive for MAP2K4 but negative for vimentin had the longest survival time, while those negative for MAP2K4 and positive for vimentin had lowest survival rate (P=0.004). CONCLUSION: Detection of MAP2K4 and vimentin might help in early diagnosis and prognostic evaluation of patients with EC.

An unusual case of anaplastic lymphoma kinase-positive large B-cell lymphoma in an elderly patient: A case report and discussion.

We present an unusual case of anaplastic lymphoma kinase (ALK)-positive large B-cell lymphoma, with rapid clinical progression, which occurred in a 90-year-old male patient. The patient presented with numerous enlarged lymph nodes in the neck and mediastinum. Histopathological analysis of a single lymph node detected diffuse large immunoblastic- or plasmablastic-like tumor cells, which were strongly immunoreactive for ALK in a granular cytoplasmic distribution, but negative for the expression of CD20 and CD79a. In addition, polymerase chain reaction assays were unable to detect clonal rearrangements of the T cell receptor-γ and immunoglobulin heavy chain genes in the tumor lesion, and in situ hybridization tested negative for infection with Epstein-Barr virus. The patient underwent a single cycle of chemotherapy using the cyclophosphamide, doxorubicin, vincristine, prednisone, and etoposide (E-CHOP) regimen; however, the patient developed pleural effusions with respiratory distress, associated with clinical deterioration. The patient succumbed to the disease within 4 months of initial presentation. To the best of our knowledge, this is the eldest patient with this type of lymphoma to be reported in the literature.

Influence of gonadotropin-releasing hormone agonist on the effect of chemotherapy upon ovarian cancer and the prevention of chemotherapy-induced ovarian damage: an experimental study with nu/nu athymic mice.

BACKGROUND AND OBJECTIVE: Gonadotropin-releasing hormone (GnRH) plays an important role in the regulation of ovarian function and ovarian cancer cell growth. In this study, we determined whether administration of the GnRH agonist (GnRHa), triporelin, prior to cisplatin treatment affects cisplatin and/or prevents cisplatin-induced ovarian damage. METHODS: nu/nu mice were injected with ovarian cancer OVCAR-3 cells intraperitoneally. After two weeks, the mice were treated with saline (control), cisplatin, GnRHa, or cisplatin plus GnRHa for four weeks. At the end of the experimental protocol, blood, tumor, ovary, and uterine tissues were resected for hematoxylin and eosin (H&E) staining, immunohistochemical analyses of Ki67, nuclear factor-κB (NF-κB), and caspase-3, transmission electron microscopy of apoptosis, or enzyme-linked immunosorbent assay (ELISA) analyses of anti-Mullerian hormone (AMH). RESULTS: Cisplatin treatment effectively inhibited tumor growth in mice treated with human ovarian cancer cells; however the treatment also induced considerable toxicity. Immunohistochemical analyses showed that Ki67 expression was reduced in cisplatin-treated mice compared to control (P<0.05), but there was no statistically significant differences between cisplatin-treated mice and cisplatin plus GnRHa-treated mice (P>0.05), while expressions of NF-κB and caspase-3 were reduced and induced, respectively, in cisplatin-treated mice and cisplatin plus GnRHa-treated mice. Apoptosis occurred in the GnRHa, cisplatin, and cisplatin plus GnRHa-treated mice, but not in control mice. Ovaries exposed to GnRHa in both GnRHa mice and cisplatin-treated mice (combination group) had significantly more primordial and growth follicles and serum levels of AMH than those in the control mice and cisplatin-treated mice (P<0.05). CONCLUSIONS: Administration of GnRHa to mice significantly decreased the extent of ovarian damage induced by cisplatin, but did not affect the anti-tumor activity of cisplatin.

CD20-positive NK/T-cell lymphoma with indolent clinical course: report of case and review of literature.

CD20-positive T-cell lymphoma is extremely rare and only two cases of CD20-positive NK/T-cell lymphoma with aggressive clinical courses have been described in the literature. We present a case of unusual NK/T-cell lymphoma with CD20 expression in nasal cavity occurring in an elder female patient. The patient had presented with left nasal cavity nodule for 10 years. CT scan revealed a mass was located at the left anterior nasal cavity and was observed to extend into the ethmoid sinus. There was no regional lymph node involvement. Biopsy was performed and microscopical inspection revealed the lesion was composed of small- to middle-size atypical lymphoid cell, histiocytes, eosinophils, and neutrophils. The lymphoid cells were strongly immunoreactive to CD3, CD20, CD56, TIA-1 and granzyme-B. The Epstein-Barr virus genomes were also found in tumor cells by in situ hybridization. By genetic analysis, however, no clonal rearrangement of the T cell receptor-γ genes (TCRG), or the immunoglobulin heavy chain (IgH) gene was found. A diagnosis of CD20-positive extranodal NK/T-cell lymphoma, nasal type was made. The patient refused chemotherapy, and had been only on regular follow-up for 6 months. There was no sign of enlargement of tumor and extra-nasal dissemination by whole body positron emission tomography/computed tomography (PET/CT) study. The accurate diagnosis of NK/T-cell lymphoma with CD20 expression is important, but the indolent behavior of the present case is more unusual. A long-term follow-up is suggested to be performed to inspect the progression for this tumor. VIRTUAL SLIDES: The virtual slides for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1320848277788495.

[Construction of a lentiviral vector containing human NESG1 gene and its expression in 293FT cells].

OBJECTIVE: To construct a lentiviral vector carrying human NESG1-EGFP gene and observe its expression in 293FT cells. METHODS: The CDS region of NESG1 gene was amplified from a plasmid containing the full-length NESG1 sequence and cloned into the lentiviral vector pGC-FU-EGFP by restriction endonuclease AgeI digestion and T(4) DNA ligase ligation. After transformation into competent E. coli cells, the candidate clones were identified by PCR and sequencing. The recombinant plasmid and the two packaging plasmids were co-transfected into human embryonic kidney cell line 293FT cells by lipofectamine 2000 to produce the lentiviral particles, and the viral titer was determined. The 293FT cells were infected by the lentiviral particles obtained and the transfection efficiency was assessed under fluorescent microscope. Western blotting was used to detect the expression of NESG1 protein in the transfected cells. RESULTS: The lentiviral vector pGC-FU-NESG1-EGFP for NESG1 gene was constructed successfully. Strong green fluorescence was observed in 293FT cells under fluorescent microscope after co-transfection of the cells with the 3 plasmids of lentiviral vector. The virus in the supernatant reached a titer of 2×10(7) TU/ml. The transfection efficiency of the collected virus exceeded 90% in 293FT cells with a multiplicity of infection of 1. Western blotting identified the presence of NESG1 expression in the transfected 293FT cells. CONCLUSION: The lentiviral vector for NESG1 has been successfully constructed with a high yield of lentivirus, which facilitate further investigation of the roles of NESG1 gene in the development and progression of nasopharyngeal carcinoma.

[Relationship between MAP3K5 and Epstein-Barr virus-encoded miR-BART22 expression in nasopharyngeal carcinoma].

OBJECTIVE: To detect the expression of MAP3K5 and miR-BART22 encoded by Epstein-Barr virus and explore their relationship in nasopharyngeal carcinomas (NPCs). METHODS: Fifty-three archived specimens of NPCs and 30 nasopharyngitis specimens were collected for detecting the expression of EBERs and miR-BART22 by in situ hybridization, and the expression of MAP3K5 was detected using immunohistochemistry. Ten fresh NPC and 10 fresh nasopharyngitis specimens were also obtained for determining the protein expression of MAP3K5 by Western blotting. RESULTS: EBERs were positive in all the 53 NPC specimens, and miR-BART22 was positive in 49 specimens; all the 30 nasopharyngitis specimens were negative for EBER or miR-BART22. In the 53 NPC tissues, 50 were negative for MAP3K5 expression in the cancer areas but positive in the adjacent mucosal areas, with the other 3 specimens showing a weak positivity (+). In the 30 nasopharyngitis specimens, 25 showed strong MAP3K5 positivity, 3 showed weak positivity and 2 were negative for MAP3K5 (P<0.001). Western blotting showed that the expression of MAP3K5 protein was significantly higher in nasopharyngitis than in NPC tissues (P=0.029). The expression of MAP3K5 and miR-BART22 was inversely correlated (P<0.001). CONCLUSION: Compared with the adjacent mucosal tissues, NPC tissues have a lower expression of MAP3K5 but a higher expression of miR-BART22. The expression of MAP3K5 and miR-BART22 is inversely correlated, suggesting the possibility of MAP3K5 to serve as target gene of EBV miR-BART22. miR-BART22 may inhibit the expression of MAP3K5, thus reducing the protein phosphorylation of MAPK pathway downstream genes, inhibiting NPC cell apoptosis, preventing their differentiation and promoting their escape from immune surveillance.

[Dynamic contrast-enhanced CT patterns and pathologies of noncalcified pulmonary tuberculomas].

OBJECTIVE: To analyze characteristic CT enhancement patterns of noncalcified pulmonary tuberculomas and their pathological basis. METHOD: Fifty-six patients with noncalcified pulmonary tuberculomas underwent surgical resection of the tuberculomas. Enhanced CT images of these tuberculomas were reviewed and analyzed in relation to the histological findings. RESULTS: Of the 56 patients, 45 showed no enhancement in the tuberculomas, which were histologically characterized by central caseous necrosis and a poorly vascularized peripheral fibrotic zone. Eleven patients showed ring-like or eggshell enhancement, and the central low density region was histologically confirmed to be caused by caseous or liquefied necrosis, while the ring enhancement resulted pathologically from moderately or well vascularized peripheral fibrotic or granulomatous tissues. CONCLUSIONS: Pulmonary tuberculomas consists mainly of caseous necrotic tissues characterized by no enhancement and ring or eggshell enhancement on dynamic contrast-enhanced CT.

[Construction and identification of a recombinant lentivirus vector of latent membrane protein 1].

OBJECTIVE: To construct a recombinant lentivirus vector of latent membrane protein 1 (LMP1) and detect the expression of LMP1 in vitro. METHODS: The LMP1 fragment including all the exons was amplified by PCR and inserted to the downstream of CMV promoter in the lentivirus vector pCDF. The three plasmids (packaging plasmid pFIV-34N, envelope plasmid pVSV-G and target plasmid pCDF-LMP1) were packaged into 293FT cells via liposome. The virus supernatant was harvested, concentrated and titrated. Mouse B lymphoma cell line A20 was transfected with the recombinant lentivirus vector of LMP1, and the expression of LMP1 in A20 cells was detected by RT-PCR and Western blotting. RESULTS: DNA sequencing confirmed that the sequence of PCR-amplified LMP1 was consistent with the GenBank data. The LMP1 gene fragment was cloned into pCDF in the right direction, and the open reading frame of LMP1 was maintained. The 3 plasmids were effectively transferred into 293FT cells, which emitted green fluorescence in the cytoplasm and on the cell membrane under fluorescence microscope. The titer of the lentivirus vector reached 10(7) Tu/ml with a transfection efficiency 90% in A20 cells. LMP1 expression was detected by RT-PCR and Western blotting in transfected A20 cells. CONCLUSION: The recombinant lentivirus vector of LMP1 constructed can be effectively transfected into A20 cells, which provides a basis for exploring the role of LMP1 in the pathogenesis of lymphoma.