Objective To construct a prokaryotic plasmid to express the testis development related gene 1 (TDRG1) recombinant protein and obtain anti-TDRG1 mAb by immunizing mice, and to identify the biological properties of the mAb. Methods The coding sequence of TDRG1 was amplified by RT-PCR from normal human testis tissue and cloned into the vector pET21, and then was expressed in the E. coli BL 21(DE3) to get TDRG1 recombinant protein. The purified TDRG1 recombinant protein was injected to immunize the BALB/C mice to develop anti-TDRG1 mAb. Splenocytes of the immunized mice were collected and fused with the mouse myeloma cell line Sp2/0 cells. The hybridoma cells that secreted anti-TDRG1 mAb were subcloned with limited dilution. Enzyme-linked immunosorbent assay (ELISA) was used to evaluate titers and subtypes of mAb. Western blot and immunohistochemistry were used to detect specificity of mAb.Results The prokaryotic plasmid expressing the recombinant protein was constructed, and the TDRG1 recombinant protein was expressed and purified. Two hybridoma cell lines that secreted anti-TDRG1 mAb were obtained. The titer of the mAb in ascites was 1∶1.6×10~6, and the subtype of the mAb was IgG_1. Westem blot and immunohistochemistry analysis indicated the mAb showed specific combination with TDRG1 protein in human testes.Conclusion The TDRG1 recombinant protein is highly purified and has strong antigenicity. The anti-TDRG1 mAb is produced successfully.

OBJECTIVE: To determine the diagnostic value of real-time Rigiscan monitoring and Doppler ultrasonography following intracavernous injection of prostaglandin E1in erectile dysfunction(ED). METHODS: From January 1998 to December 2007, 1 392 ED patients completed the test. PGE(1) was injected into corpus cavernosum penis near the radix penis with injection guns. After 1 or 2 minutes, the Rigiscan Plus device was used for real-time evaluation of penile rigidity. Optimal injected dosage of each patient with positive reaction was determined. Injected dosage of patients with negative reaction was gradually increased from to 60 microg. As the dosage was established, color Doppler ultrasonography was used to assess the morphodynamic features of cavernosal arteries. RESULTS: Altogether 1 039 patients were positive, while 353 patients were negative by Rigiscan. In the 1 039 positive patients,663 were injected with 10 microg PGE(1), 265 patients 20 microg, 97 patients 30 microg,and 14 patients 60 microg while 89(13.4%),39(14.7%),41(42.3%),and 10(71.4%) patients were diagnosed vascular ED. Of the 353 negative patients, 267(75.64%) patients were diagnosed vascular ED. During intracavernous pharmacological testing,the more PGE(1) was used, the more vascular ED patients would be found(P<0.001). CONCLUSION PGE(1) is safe and effective for the erectile responses. The dosage of PGE(1) following intracavernous injection is related to vascular ED. Real-time Rigiscan monitoring combined with color Doppler ultrasonograph can be helpful to diagnose penile vascular diseases.
Adult ; Alprostadil ; administration & dosage ; Erectile Dysfunction ; diagnosis ; diagnostic imaging ; physiopathology ; Humans ; Male ; Penile Erection ; Penis ; blood supply ; diagnostic imaging ; Ultrasonography, Doppler, Color

OBJECTIVE: To construct short hairpin RNA interfering expression vector of TDRG1,and detect the specific interfering effect of TDRG1-shRNA expression vector on NTERA-2 cells. METHODS: Oligos for short hairpin RNA targefing for TDRG1 were designed and connected to the expression vector pGPU6/GFP/Neo to construct the TDRG1 shRNA expression vector. The recombinant plasmid TDRG1-shRNA486, TDRG1-shRNA738, TDRG1-shRNA921 and lipofectamine ™2000 were used to generate and transfect shRNA into NTERA-2 cells. Expression of TDRG1 mRNA was assayed by RT-PCR. RESULTS: TDRG1-shRNA expression vector was successfully constructed. TDRG1-shRNA486 was more effective in the suppression of TDRG1 with significant reduction of TDRG1 mRNA. CONCLUSION TDRG1-shRNA can interfere the expression of TDRG1 in NTERA-2 cells.
Cell Line, Tumor ; Genetic Vectors ; Humans ; RNA Interference ; RNA, Messenger ; RNA, Small Interfering ; Transfection

Objective:To analyze the dynamic changes of T lymphocytes in patients with COVID-19.Methods:Blood samples were collected from 40 COVID-19 cases and 40 healthy controls in Beihai People′s Hospital from January to February, 2020. The counts of CD4 + T and CD8 + T lymphocytes were detected by flow cytometry. Moreover, the T lymphocyte counts in 24 convalescent patients with two consecutive negative nucleic acid test results were also detected. Results:The leukocytes and lymphocytes in the patients with acute COVID-19 were significantly lower than those in the healthy controls [(4.71±1.54)×10 9 cell/L vs (6.26±1.44)×10 9 cell/L, (1.13±0.41)×10 9 cell/L vs (1.51±0.39)×10 9 cell/L; both P<0.05]. The counts of CD4 + T and CD8 + T lymphocytes in the patients with acute COVID-19 were significantly lower than those in the healthy controls [(447.15±144.42) cell/μl vs (592.83±146.76) cell/μl, (309.35±173.05) cell/μl vs (397.20±136.94) cell/μl; both P<0.05], while no significant difference was observed in the CD4 + /CD8 + T cell ratio ( P>0.05). In the 24 convalescent COVID-19 patients, the counts of CD4 + T and CD8 + T lymphocytes were higher during convalescence than in the acute phase [(598.08±138.71) cell/μl vs (420.67±147.38) cell/μl, (439.08±166.94) cell/μl vs (296.67±151.06) cell/μl; both P<0.05], but there was no significant difference in the T lymphocyte counts between the convalescent patients and the healthy controls ( P>0.05). Conclusions:A transient immune deficiency occurred in patients with acute COVID-19, but the impaired immune function could restore to normal level during recovery.