Associations of serum cryptococcal antigen with different of clinical characteristics: a comprehensive analysis of 378 pulmonary cryptococcosis patients.

BACKGROUND: Pulmonary cryptococcosis (PC) is an infection typically diagnosed in immunocompromised or immunocompetent patients, which can lead to severe disease if not treated appropriately. We aimed to determine the association between clinical manifestations, computed tomography (CT) findings, and host immune status with the serum cryptococcal antigen (CRAG) test results of PC patients. METHODS: The clinical data of 378 PC patients over a 12-year period were retrospectively reviewed at Shanghai Pulmonary Hospital (Shanghai, China). Serum CRAG was detected by a latex agglutination (LA) test using CryptoTrol (Immuno-Mycologics Inc., Norman, OK, USA). Patients were categorized according to their serum LA results, and their clinical characteristics were analyzed: 244 of 378 patients showed positive serum LA results and 134 had negative results. RESULTS: Immunocompromised hosts (ICH) were more likely to present positive LA results. The ICH group had higher titers of LA test than the non-immunocompromised host (NICH) group. Patients with negative LA results often had no symptoms and their CT findings presented a solitary nodule or mass, while LA-positive patients had variable symptoms such as cough, expectoration, fever, etc. A large diversity of CT manifestations were observed in the LA-positive patients, such as multiple nodules, patchy shadows, interstitial infiltrates, and diffuse granular shadows. Patients with a solitary nodule or mass had lower titers than did the patients with other manifestations. The clinical characteristics of LA-positive patients were different from those of LA-negative patients. CONCLUSIONS: Serum CRAG test results were found to be associated with the clinical manifestations, CT findings, and host immune status of PC patients.

[Impacts of Chk1 and Chk2 gene expressions on sperm concentration and motility].

OBJECTIVE: To study the correlation of the gene expressions of Chk1 and Chk2 with sperm concentration and motility. METHODS: According to sperm concentration and motility (percentage of progressively motile sperm), we divided 80 semen samples into four groups of equal number: normal control, oligozoospermia (OS), asthenospermia (AS), and oligoasthenozoospermia (OAS). We detected the sperm DNA fragmentation index (DFI) and viability and determined the expressions of Chk1 and Chk2 in the sperm by RT-PCR and Western blot. RESULTS: Statistically significant differences were not found in sperm DFI among the control, OS, AS, and OAS groups (21.24±6.93, 19.67±7.64, 21.52±6.92, and 19.28±11.55, P>0.05), but observed in sperm concentration, progressive motility, and viability between the DFI >30% and DFI ≤30% groups (P<0.01). Compared with the normal control, sperm viability was remarkably decreased in the OS, AS, and OAS groups (ï¼»83.48±9.87ï¼½% vs ï¼»63.86±9.16ï¼½%, ï¼»50.45±16.99ï¼½%, and ï¼»39.21±15.74ï¼½%, P<0.05). RT-PCR showed remarkable differences among the control, OS, AS, and OAS groups in the relative expression level of Chk1 mRNA (0.73±0.22, 0.62±0.14, 1.03±0.39, and 0.92±0.071, P<0.01), which was correlated positively with sperm concentration (b = 80.661, P<0.01) but negatively with sperm motility (b = －19.275, P < 0.01), as well as in that of Chk2 mRNA (0.66±0.30, 0.27±0.09, 0.59±0.19, and 0.42 ± 0.11, P<0.01), which was correlated negatively with sperm concentration (b = －90.809, P<0.01) but positively with sperm motility (b = 27.507, P <0.01). The relative expression levels of the Chk1 protein were significantly different among the four groups (0.63±0.05, 0.42±0.03, 1.13±0.08, and 0.87±0.07, P<0.01), which was correlated positively with sperm concentration (b = 55.74, P<0.01) but negatively with sperm motility (b =－22.649, P<0.01), and so were those of the Chk2 protein (1.23±0.36, 0.37±0.16, 0.87±0.08, and 0.68±0.12, P<0.01), which was correlated negatively with sperm concentration (b =－53.001, P<0.01) but positively with sperm motility (b = 16.676, P < 0.01). CONCLUSIONS: Chk1 and Chk2 are significantly expressed in human sperm. In case of sperm DNA damage, up-regulated Chk1 expression may enhance sperm apoptosis and lead to asthenospermia, while increased Chk2 expression may inhibit spermatogenesis and result in oligospermia.

Glucose regulated protein 78 (GRP78) inhibits apoptosis and attentinutes chemosensitivity of gemcitabine in breast cancer cell via AKT/mitochondrial apoptotic pathway.

The underlying mechanism of gemcitabine resistance during breast cancer treatment remains unclear. Glucose regulated protein 78 (GRP78) frequently triggered by anticancer agents, was substantially elevated in gemcitabine resistant sublines. Ectopic expression of GRP78 changes gemcitabine chemosensitivity and apoptosis levels in breast cancer cells. Further experiments showed an involvement of caspase 9, not caspase 8, in gemcitabine resistance and GRP78-mediated chemosensitivity, suggesting that mitochondria apoptotic pathway was activated by GRP78. This finding was further supported by the observations of AKT activation, Bcl-2 increase, Bax and Bim decrease. Conclusively, GRP78 plays a vital role in gemcitabine resistance and clinical strategy to improve gemcitabine efficacy in breast cancer by manipulating GRP78 should be explored.

[Tea polyphenols reduces the apoptosis of spermatogenic cells in rats with experimental varicocele].

OBJECTIVE: To study the effect of tea polyphenols (TP) on the apoptosis of germ cells in rats with experimental varicocele. METHODS: Thirty-two adolescent male Wistar rats were randomly and equally divided into groups A (sham-operation), B (high-dose TP), C (low-dose TP), and D (experimental left varicocele). Experimental varicocele was induced by partial ligation of the left renal vein in the latter three groups of rats. The animals in groups A and D were fed with normal saline, while those in B and C with TP at 40 and 10 mg per kg per d, respectively, all for 4 weeks. Then, all the rats were sacrificed and the left testes harvested for determination of the expression of HIF-1, Bcl-2, Bax, CytC, and caspase-3 by immunohistochemistry and measurement of the apoptosis index (AI) of spermatogenic cells. RESULTS: The expression of Bcl-2 was higher in groups B and C than in D but lower than in A (P < 0.05), and lower in C than in B (P < 0.05). However, the expressions of HIF-1, Bax, CytC, and caspase-3 were lower in groups B and C than in D but higher than in A (P < 0.05), and higher in C than in B (P < 0.05). The AI of spermatogenic cells was the lowest in group A, higher in D than in the other groups but lower in B than in C (P < 0.05). CONCLUSION: TP can reduce the apoptosis of spermatogenic cells in a dose-dependent manner in varicocele rats.

[Effect of different porcelain-fused-to-metal crown inner metal materials on the level of soluble intercellular adhesion molecule-1 and interleukin-1beta in gingival crevicular fluid].

OBJECTIVE: To investigate the effect of different inner metal materials of porcelain-fused-to-metal (PFM) crown on periodontal tissue by means of measuring the level of soluble intercellular adhesion molecule-1 (sICAM-1) and interleukin-1beta (IL-1beta) in gingival crevicular fluid (GCF) after PFM restorations. METHODS: 30 teeth were divided into three groups (Ni-Cr alloy group, Co-Cr alloy group and Au-Pt alloy group, 10 teeth each group), and restored by Ni-Cr alloy, Co-Cr alloy and Au-Pt alloy PFM crown according grouping. At the point of pre-restoration, 6-month and 12-month after cementation, the clinical parameters including plaque index (PLI), gingival index (GI) and gingival crevice depth (GCD) were detected, and GCF was collected from labial and lingual of mesial site and distal site. The level of sICAM-1 and IL-1beta were detected. RESULTS: At the point of 6-month and 12-month after cementation, Ni-Cr alloy group showed significant difference for GI, GCD and all GCF indexes when compared to pre-restoration, Co-Cr alloy group and Au-Pt alloy group (P < 0.05). At the point of 12-month after cementation, Co-Cr alloy group showed significant difference for GI, GCD and all GCF indexes when compared to pre-restoration and Au-Pt alloy group (P < 0.05). All indexes have no significant difference for Au-Pt alloy group during the 12-month experiment times when compared to pre-restoration (P > 0.05). CONCLUSION: Non-noble metal has bad effect on the periodontal tissue.

[Preparation, characterization and potential application of monoclonal antibody 18A4 against AGR2].

AIM: To prepare mouse monoclonal antibodies (mAb) against AGR2 (human homolog of xenopus anterior gradient 2), to characterize these antibodies' properties, and to develop potential applications. METHODS: BALB/c mice were immunized with AGR2-MBP (maltose binding protein) fusion protein. The mAb was prepared by hybridoma technique and purified by protein G affinity chromatography. The titer and specificity of the mAb was determined by ELISA and Western blot respectively. The mAb was then further characterized by immunoprecipitation, immunofluorescent staining and tumor cell inhibition assay. RESULTS: One clone of hybridoma, 18A4, secreting specific mAb against AGR2, was obtained. The Ig subclass of the mAb was IgG1 (kappa). The titer of the mAb was 1 x 10(-6);. Western blot analysis showed specific binding of 18A4 with both recombinant and native AGR2 in cell extract. Immunofluorescent staining using 18A4 mAb demonstrated specific staining of AGR2 in the cytoplasma of Breast cancer cell line MCF7. Immunoprecipitation assay confirmed that this mAb could specifically bind and effectively precipitate the native AGR2. mAb 18A4 could also inhibit the growth of breast cancer cell MCF7. CONCLUSION: The mAb anti-AGR2 with high titer and specificity has been obtained. This mAb, 18A4, can be used in most molecular biology studies including Western blot, immunostaining, immunoprecipitation. It also inhibited the growth of cancer cells and therefore is a potential therapeutic starting point. It is a useful tool for the functional study of AGR2 and for the diagnosis and potential treatment of certain cancers.

Apoptosis of murine melanoma cells induced by heavy-ion radiation combined with Tp53 gene transfer.

PURPOSE: To investigate the effect of exogenous wild type p53 (Tp53) on murine melanoma B16 cell apoptosis induced by carbon-ion beam (C-beam) irradiation. MATERIALS AND METHODS: The murine cell line B16, which has wild-type Tp53 gene status was studied, as well as B16 cells transfected with an adenoviral vector containing the wild-type Tp53 gene (B16/Tp53). Cells were irradiated with C-beam and assayed for cell survival (colony-forming assay), cellular morphology (acridine orange assay), the frequency of apoptotic cells (fluorescence microscopy) and protein expression (Western blot analysis). RESULTS: The radiosensitivity of B16/Tp53 cells was significantly higher than that of B16 cells. In contrast with Tp53 transfer alone, the combination of C-beam with Tp53 transfer induced a higher proportion of apoptotic cells and micronuclei. After C-beam irradiation, there was no significant increased expression of the cyclin-dependent kinase inhibitor p21 and Tp53 in B16/Tp53 cells compared to B16 cells, but a decreased expression of murine double minute-2 (Mdm2) was observed. CONCLUSION: The results of this study suggest the potential application of C-beam combined with Tp53 in the treatment of melanoma in human patients.

[Preparation and characterization of monoclonal antibody against human carboxylesterases-II].

AIM: To prepare monoclonal antibody(mAb) against human carboxylesterases-II (hCE-II) and characterize its properties. METHODS: BALB/c mice were immunized with human liver microsome protein which contained hCE-II. The mAb was prepared by hybridoma technique and purified by protein-G affinity chromatography. The titer and specificity of mAb was detected by ELISA and Western blot respectively. Tissue localization of antigen was detected by immunohistochemical staining. Antigen was appraised by peptide mass fingerprint (PMF) matched with Mascot human protein database. PMF was obtained by immunoprecipitation and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). RESULTS: One clone of hybridoma secreting specific mAb against hCE-II was obtained. The Ig subclass of the mAb was IgG1(kappa). The titer of the mAb was 1 x 10(-7). Western blot analysis showed one clear belt in the Mr of 62,000. Immunohistochemistry demonstrated that the mAb had special combination with the liver cytoplasm protein, but not with the vascular smooth muscle cell protein. Immunoprecipitation showed one clear band in the Mr of 62,000, which was in conformity with the Mr of hCE-II and the antigen was confirmed to be hCE-II after being analyzed with mass spectrometry. CONCLUSION: The mAb against hCE-II with high titer and specificity has been obtained, which lays the foundation for investigation of hCE-II function and diagnosis and therapy of liver cancer.

[Multivariate analysis of impact of different neck treatment and prognostic factors affecting disease-free survival in patients with early-staged squamous cell carcinoma of tongue].

OBJECTIVE: To analyze the prognostic factors affecting the disease-free survival in T1/T2N0M0-staged patients with squamous cell carcinoma of tongue and compare the effectiveness of different neck treatment modalities. METHODS: 97 consecutive patients with early-staged squamous cell carcinoma of tongue were included in this study. The treatment and following-up records were reviewed retrospectively. Cox proportional hazard model was used to identify the statistically significant prognostic factors in the 6 potential factors. Kaplan-Meier method was used to estimate the disease-free survival and analyze the survival rate among the different levels, and log-rank method for comparison of the different distribution of the survival. A special focus was on the effectiveness of different neck treatment modalities. RESULTS: T stage, treatment methods of primary tumor, the modalities of neck treatment and cell differentiation were statistically significant prognostic factors. The value of P and relative risk (RR) were P < 0.001, RR = 4.387; P = 0.04, RR = 0.496; P = 0.003, RR = 0.504; P < 0.001, RR = 2.620, respectively. The difference of disease-free survival was statistically significant among the different levels under the different factors. CONCLUSION: The disease-free survival was affected by neck treatment modalities remarkably in cN0 stage patients. Selected neck dissection together with adjuvant irradiation could decrease the recurrence risk by 49.6% according to the results of this study. TNM stage system could describe the characteristics of the patients with early-staged squamous cell carcinoma of tongue reasonably.

[Experimental studies on ectopic osteogenesis by seeding human mesenchymal stem cell on human natural bone derived scaffold materials].

OBJECTIVE: To investigate the ectopic osteogenesis potential of human natural bone derived material combined with human bone marrow mesenchymal stem cells (MSCs). METHODS: Cell-scaffold complexes were implanted subcutaneously into the left back of the nude mice, and human natural bone derived material were implanted into the right back as control group. The mice were killed respectively on the postoperative 2 weeks, 4 weeks and 8 weeks. The macroscopic, histopathological, alkaline phosphatase (ALP) activity assay methods were performed to assess the ectopic osteogenesis potential. RESULTS: The cartilaginous osteogenesis were observed in both deproteinated bone and decalcified bone, and the more new bone tissue formed gradually as the time went by after implantation. ALP activity become stronger followed with the time (P < 0.05), and compared with the decalcified bone, deproteinated bone displayed stronger ALP activity (P < 0.05). CONCLUSION: The MSCs and human natural bone derived material can be used as good seed cells and scaffold materials respectively to construct tissue-engineered bone, and as the scaffold material, deproteinated bone has better osteogenesis ability than decalcifed bone.