Identification of clinical prognostic factors and analysis of ferroptosis-related gene signatures in the bladder cancer immune microenvironment.

BACKGROUND: Bladder cancer (BLCA) is a prevalent malignancy affecting the urinary system and poses a significant burden in terms of both incidence and mortality rates on a global scale. Among all BLCA cases, non-muscle invasive bladder cancer constitutes approximately 75% of the total. In recent years, the concept of ferroptosis, an iron-dependent form of regulated cell death marked by the accumulation of lipid peroxides, has captured the attention of researchers worldwide. Nevertheless, the precise involvement of ferroptosis-related genes (FRGs) in the anti-BLCA response remains inadequately elucidated. METHODS: The integration of BLCA samples from the TCGA and GEO datasets facilitated the quantitative evaluation of FRGs, offering potential insights into their predictive capabilities. Leveraging the wealth of information encompassing mRNAsi, gene mutations, CNV, TMB, and clinical features within these datasets further enriched the analysis, augmenting its robustness and reliability. Through the utilization of Lasso regression, a prediction model was developed, enabling accurate prognostic assessments within the context of BLCA. Additionally, co-expression analysis shed light on the complex relationship between gene expression patterns and FRGs, unraveling their functional relevance and potential implications in BLCA. RESULTS: FRGs exhibited increased expression levels in the high-risk cohort of BLCA patients, even in the absence of other clinical indicators, suggesting their potential as prognostic markers. GSEA revealed enrichment of immunological and tumor-related pathways specifically in the high-risk group. Furthermore, notable differences were observed in immune function and m6a gene expression between the low- and high-risk groups. Several genes, including MYBPH, SOST, SPRR2A, and CRNN, were found to potentially participate in the oncogenic processes underlying BLCA. Additionally, CYP4F8, PDZD3, CRTAC1, and LRTM1 were identified as potential tumor suppressor genes. Significant discrepancies in immunological function and m6a gene expression were observed between the two risk groups, further highlighting the distinct molecular characteristics associated with different prognostic outcomes. Notably, strong correlations were observed among the prognostic model, CNVs, SNPs, and drug sensitivity profiles. CONCLUSIONS: FRGs are associated with the onset and progression of BLCA. A FRGs signature offers a viable alternative to predict BLCA, and these FRGs show a prospective research area for BLCA targeted treatment in the future.

Quality Care Alleviates Behavioral Cognitive Impairment and Reduces Complications in Elderly Patients with Cardiovascular and Cerebrovascular Diseases.

Objective: Cardiovascular and cerebrovascular disease (CCVD) remains the most common factor of death around the world. Nursing care plays a key role in the recovery of patients with CCVD. This study was to explore the application of quality care in aged patients with CCVD. Methods: Totally, 74 aged CCVD patients admitted from June 2018 to June 2019 in Dongying People's Hospital were randomly assigned in 2 groups with the same treatment. The control group was treated with routine care intervention, and the observation group was treated with quality care intervention for 12 weeks. Meanwhile, the frequency of agitation behaviors and cognitive ability were assessed, and complication was counted. Results: The observation group showed decreased Cohen-Mansfield Agitation Inventory (CAMI) scores from 47.31 ± 8.27 to 38.73 ± 6.94, raised Mini-Mental State Examination (MMSE) scores from 15.01 ± 3.9 to 19.34 ± 3.15 and Montreal Cognitive Assessment (MoCA) scores from 16.92 ± 5.48 to 20.37 ± 4.16, and reduced complications after quality care intervention. Conclusion: Quality care intervention exerted a better application effect on aged CCVD patients, along with reduction of agitation, improvement of mental condition and behavioral cognitive function, and reduced complications.

Protective effect of hypothermia and vitamin E on spermatogenic function after reduction of testicular torsion in rats.

This study was designed to investigate the protective effect of hypothermia and vitamin E on spermatogenic function after reduction of testicular torsion in rats. Ninety-six pure inbred male SD rats were divided into group A, B, C and D according to the principle of body weight and birth similarity, with 24 rats in each group. Four groups of rats were respectively twisted on the left testis to establish unilateral testicular torsion rats. Rats in groups A, B, C, D were respectively given normal saline, hypothermia therapy, vitamin E therapy, and hypothermia and vitamin E therapy. The superoxide dismutase (SOD) activity and malondialdehyde (MDA) content of the four groups were detected, and the correlation levels of inflammatory factors IL-1ß, hs-CRP and related sex hormones luteinizing hormone (LH), follicle-stimulating hormone (FSH), total testosterone (T) were detected by ELISA. Apoptosis of spermatogenic cells of testis in the four groups was detected by flow cytometry. SOD activity and MDA content in groups B, C and D were significantly higher than those in group A, MDA content was significantly lower than that in group A (P<0.05), SOD activity in group D was higher than that in groups B and C, while MDA content was lower than that in groups B and C (P<0.05). The levels of IL-1ß and hs-CRP in group A were much higher than those in groups B, C and D (P<0.05). LH and FSH levels in group A were significantly higher than those in groups B, C and D (P<0.05), and in group D were significantly lower than those in groups B and C (P<0.05). Apoptosis rate of spermatogenic cells in group A was significantly higher than that in groups B, C and D (P<0.05). Hypothermia combined with vitamin E can reverse testicular injury in rats and reduce the apoptosis rate of spermatogenic cells.

sBAFF mutants induce neutralizing antibodies against BAFF.

B cell activating factor belonging to the TNF family (BAFF) is a novel member of the tumor necrosis factor (TNF) ligand family and plays an important role in B lymphocyte maturation and survival. Overexpression of BAFF is closely involved in the pathogenesis and progression of many kinds of autoimmune disorders; therefore, BAFF has been considered as an ideal therapeutic target for these conditions. In this study, we generated several candidate immune inhibitors of human BAFF by conjugating foreign immunodominant T-helper cell (Th) epitopes to the N- or C-terminus of five BAFF mutants. The recombined proteins were successfully expressed in Escherichia coli (E. coli) and purified by Ni-NTA chromatography. BALB/c mice immunized with the recombinant proteins produced high levels of anti-BAFF antibodies, and their sera inhibited the lymphocyte proliferation-inducing activity of recombinant soluble BAFF and natural soluble BAFF. Moreover, antibodies cross-reactive with BAFF were detected in sera from hu-SCID mice immunized with the recombinant proteins. These results indicated that the recombinant BAFF mutants modified with Th epitopes could induce neutralizing antibodies against BAFF in vivo. This study may provide a valuable strategy for treating BAFF-associated autoimmune diseases.

Interleukin-1 up-regulates the expression and activity of 26S proteasome in burned rat.

Expression of IL-1 and proteasome are elevated in burned animals and patients. However, whether the increased level of IL-1 correlates with the increased activity and expression of 26S proteasome after burn has not been studied. In the present study, we investigated the role of single IL-1 factor on activation of the 26S proteasome first by injection of recombinant IL-1 into the normal rats. Results indicated that proteolytic activity and the expression of the 26S proteasome increased remarkably 24 and 48 h after-IL-1 injection, respectively. We then studied the potential role of IL-1 on activity and expression of the proteasome in the burned rat by using neutralizing monoclonal antibody against IL-1. Results demonstrated that activity and the expression of 26S proteasome were decreased partially but significantly 48 h after-burn when circulating IL-1 in injured animals was neutralized. These results indicate that IL-1 may play a key role on the activity and expression of 26S proteasome following burn. The proteasome has been verified as being deeply involved in the mechanism of accelerated muscle protein breakdown after burn, these results imply that IL-1 might be involved in the protein metabolism after-burn by activating the proteasome pathway, though protein metabolism directly affected by IL-1 had not been assessed in this study.

Imaging diagnosis of 12 patients with hepatic tuberculosis.

AIM: To assess CT, MR manifestations and their diagnostic value in hepatic tuberculosis. METHODS: CT findings in 12 cases and MR findings in 4 cases of hepatic tuberculosis proved by surgery or biopsy were retrospectively analyzed. RESULTS: (1) CT findings: One case of serohepatic type of hepatic tuberculosis had multiple-nodular lesions in the subcapsule of liver. Parenchymal type was found in 10 cases, including multiple, miliary, micronodular and low-density lesions with miliary calcifications in 2 cases; singular, low-density mass with multiple flecked calcifications in 3 cases; multiple cystic lesions in 1 case; multiple micronodular and low-density lesions fusing into multiloculated cystic mass or "cluster" sign in 3 cases; and singular, macronodular and low-density lesion with multiple miliary calcifications in 1 case. One case of tuberculous cholangitis showed marked dilated intrahepatic ducts with multiple flecked calcifications in the porta hepatis. (2) MR findings in 4 cases were hypointense on both T1-weighted imagings and T2-weighted imagings in one case, hypointense on T1-weighted imagings and hyperintense on T2-weighted imagings in 3 cases. Enhanced MR in 3 cases was slightly shown peripheral enhancement or with multilocular enhancement. CONCLUSION: Various types of hepatic tuberculosis have different imaging findings, and typical CT and MR findings can suggest the diagnosis.

Expression and identification of recombinant soluble single-chain variable fragment of monoclonal antibody MC3.

AIM: To generate soluble single chain variable fragments (ScFv) of monoclonal antibody MC3 recognizing colorectal and gastric carcinomas. METHODS: mRNA was isolated from the hybridoma cell line producing MC3 and the DNAs encoding variable domains of heavy and light chains (VH and VL) of the antibody were amplified separately by RT-PCR and assembled into ScFv DNA with a linker DNA. The ScFv DNA was ligated into the phagemid vector pCANTAB5E and the ligated sample was transformed into E.coli TG1. The transformed cells were infected with M13KO7 helper phage to yield recombinant phages. After two rounds of panning with gastric carcinoma cell line AGS highly expressing MC3-binding antigen, the phage clones displaying ScFv fragments of the antibody were selected by ELISA. 4 phage clones showing strong signal in ELISA were used to infect E.coli HB2151 to express soluble ScFvs. The soluble ScFvs were identified by Dot blot and Western blot, and their antigen-binding activity was assayed by ELISA. The VH and VL DNAs of the ScFv DNA derived from phage clone 19 were sequenced. RESULTS: The VH,VL and ScFv DNAs were about 340 bp, 320 bp and 750 bp respectively. After two rounds of panning to the recombinant phages, 18 antigen-positive phage clones were selected from 30 preselected phage clones by ELISA. All the soluble ScFvs derived from the 4 out of the 18 antigen-positive phage clones were about M(r)32000 and concentrated in periplasmatic space under the given culture condition. The soluble ScFvs could bind the antigen, and they shared the same binding site with MC3. The sequences of the VH and VL DNAs of the MC3 ScFv showed that the variable antibody genes belonged to the IgG1 subgroup,kappa-type. CONCLUSION: The soluble ScFv of MC3 is successfully produced, which not only provides a possible novel targeting vehicle for in vivo and in vitro study on associated cancers, but also offers the antibody a stable genetic source.

[CT manifestations of primary hepatic sarcoma].

OBJECTIVE: To assess the diagnostic value of CT to primary hepatic sarcoma. METHODS: The CT findings of 12 cases of primary hepatic sarcomas confirmed by operation and pathology, including leimyosarcoma, liposarcoma, and malignant fibrous histocytoma, 2 cases each, and angiosarcoma, carcinosarcoma, cystadenocarcinosarcoma, mesotheliosarcoma, fibrosarcoma, and malignant lymphoma, one case each, were analysed retrospectively. RESULTS: Only 2 out of the 12 cases of primary hepatic sarcoma were correctly diagnosed by CT before operation. The CT manifestations of primary hepatic sarcoma, against the operational and pathological findings, could be summarized as two types: solid mass type and cystic mass type. Eight cases were of solid mass type, including 2 cases of liposarcoma, and leiomyosarcoma, angiosarcoma, carcinosarcoma, methotheliosarcoma, fibrosarcoma, and malignant lymphoma, one case each. Plain scanning showed homogeneous or inhomogeneous low-density masses; enhanced scanning showed no marked enhancement or inhomogeneous enhancement, ringed peripheral enhancement, or nodular peripheral enhancement. Peripheral enhancement was a CT feature common to angiosarcoma and lymphoma. The characteristic CT manifestation of liposarcoma was an inhomogeneous mass with well-defined border and multiple intermingled septa. Four cases were of cystic mass type, including 2 cases of malignant fibrous histocytoma, and leimyosarcoma with post-operative recurrence and cystadenocarcinosarcoma, one case each, all manifesting a huge single cystic mass: unilocular in the case of leimyosarcoma with post-operative recurrence and multilocular in the other 3 cases. The wall and/or septa of the cyst had an uneven thickness and were markedly enhanced after contrast administration. Clear solid element and wall nodules were seen in the cystic mass of cystadenocarcinosarcoma. CONCLUSION: The CT findings of primary hepatic sarcoma are associated with the pathology of tumor. Some pathological patterns of primary hepatic sarcoma, such as liposarcoma and angiosarcoma, have their own characteristic CT manifestations. However, the diagnosis of most primary hepatic sarcoma lacking characteristic CT manifestations must be based on clinical and laboratory examinations and confirmed by pathology.

Expression and purification of a soluble B lymphocyte stimulator mutant modified with the T-helper cell epitope.

The DNA encoding soluble B lymphocyte stimulator (134-285 amino acids, sBLyS) mutant with residues 217-224 replaced by two glycines (named msBLyS) was constructed. The sequence encoding a foreign immunodominant T-helper epitope from ovalbumin (OVA) was then coupled to the 5'-end of msBLyS cDNA. After being sequenced, the recombinant DNA was ligated into the prokaryotic expression vector pQE-80L. The recombinant protein was produced in E. coli DH5alpha after induction with IPTG with the yield of more than 40% of total bacterial protein. The recombinant protein was purified with Ni-NTA chromatography and Sepharcryl S200 chromatography to a purity of more than 98%. The BALB/c mice, immunized with the recombinant protein, produced anti-BLyS antibodies at a high level, which indicated that the recombinant BLyS mutant modified with T-helper epitope elicited polyclonal antibodies with cross-reactivity with BLyS in vivo. This recombinant protein may therefore be used as immune inhibitor of BLyS for treating BLyS -associated autoimmune diseases.

[Preparation of phage-displayed single chain variable fragments of monoclonal antibody MC5 recognizing colorectal carcinoma].

BACKGROUND & OBJECTIVE: MC5 is a murine monoclonal antibody with a good specificity to human colorectal carcinoma and smaller murine antibody can significantly decrease the possibility of developing human antimouse antibody response in vivo study. The aim of this study was to prepare single chain variable fragments (ScFv) of MC5. METHODS: mRNA was isolated from the hybridoma cell line producing MC5, and the DNAs encoding variable domains of heavy and light chains(VH and VL DNAs) of the antibody were amplified separately by RT-PCR and assembled into ScFv DNAs with a linker DNA. The ScFv DNAs were ligated into the phagemid vector pCANTAB5E and the ligated sample was transformed into E. coli TG1. The transformed cells were infected with M13KO7 helper phage to yield recombinant phage antibody ScFvs. After two rounds of panning with cell line SW480 highly expressing MC5-binding antigen, the phage clones displayed ScFv fragments of the antibody were selected by ELISA, and the affinity of the positive phage clones was assayed by competition ELISA. RESULTS: The VH, VL, and ScFv DNAs were about 340 bp, 320 bp, and 750 bp, respectively. Ten phage clones displayed ScFv of MC5 were selected from 25 enriched phage clones, and 3 of the 10 phage clones had higher affinity of binding to the antigen. CONCLUSION: The phage-displayed ScFv fragments of monoclonal antibody MC5 are successfully produced by phage display technique, which may provide a way for broadening the application range of the antibody.

[Screening of antagonist peptide of BLyS from C7C phage display peptide library].

AIM: To screen an antagonist peptide of BLyS from C7C phage display peptide library. METHODS: C7C phage display peptide library was screened with BLyS. Indirect ELISA, competitive ELISA and MTT colorimetry were used to identify positive phage clones. RESULTS: After 3 rounds of screening, the gradual increase of the ratio of output to input and specific enrichment had been achieved. Two phage clones that could inhibit the BLyS activity were identified. CONCLUSION: Two phage display antagonist peptides of BLyS were obtained.

[The cloning, high level expression in Escherichia coli of human beta-defensin 3 and its antimicrobial activity analysis].

In recent years, Bacterial resistance is more and more serious for the irrational use of antibiotics produces resistant strains and other reasons. Human are trying to solve the problem from different ways, including the study of antimicrobial peptides. Defensin is one of the most important of antimicrobial peptides. A novel antimicrobial peptide, human beta-defensin 3, was isolated and demonstrated a salt-insensitive broad spectrum of potent antimicrobial activity against many potentially pathogenic microbes. The total RNA was extracted from human tonsil and the hbetaD-3 specific cDNA sequence was amplified with RT-PCR. After sequenced, the target DNA fragment was cloned into pQE-80L vector together with the DNA fragment encoding carrier protein DHFR. The recombinant vectors were transformed into E. coli M15 and the expression was induced based on the optimal values of the IPTG concentration incubation temperature and induction time determined in the previous section. The expressed proteins were analyzed by SDS-PAGE and Western-blotting. The mass of the recombinant protein was about 40% of total bacteria protein. Isolate and purify the target protein. The recombinant hbetaD-3 fusion proteins possess the antimicrobial activity to staphylococcus aureus, multiresistant staphylococcus aureus (only vancomycin-sensitive) and Candida albicans in the assay of drug susceptibility. Advanced study can be continued based on our experiments.