Survival after minimally invasive radical hysterectomy with protective colpotomy for early-stage cervical cancer: A systematic review and meta-analysis.

Minimally invasive surgery on treatment of early-stage cervical cancer is debatable. Traditional approaches of colpotomy are considered responsible for an inferior oncological outcome. Evidence on whether protective colpotomy could optimize minimally invasive technique and improve prognoses of women with early-stage cervical cancer remains limited. We produced a systematic review and meta-analysis to compare oncological outcomes of the patients treated by minimally invasive radical hysterectomy with protective colpotomy to those treated by open surgery according to existing literature. We explored PubMed, Embase, the Cochrane Library, and ClinicalTrials.gov from inception to December 2022. Inclusion criteria were: (1) randomized controlled trials or observational studies published in English, (2) studies comparing minimally invasive radical hysterectomy with protective colpotomy to abdominal radical hysterectomy in early-stage cervical cancer, and (3) studies comparing survival outcomes. Two reviewers performed the screening, data extraction, and quality assessment independently. A total of 8 retrospective cohort studies with 2020 women were included in the study, 821 of whom were in the minimally invasive surgery group, and 1199 of whom were in the open surgery group. The recurrence-free survival and overall survival in the minimally invasive surgery group were both similar to that in the open surgery group (pooled hazard ratio, 0.88 and 0.78, respectively; 95% confidence interval, 0.56-1.38 and 0.42-1.44, respectively). Minimally invasive radical hysterectomy with protective colpotomy on treatment of early-stage cervical cancer had similar recurrence-free survival and overall survival compared to abdominal radical hysterectomy. Protective colpotomy could be a guaranteed approach to modifying minimally invasive technique.

Co-expressions of casein kinase 2 (CK2) subunits restore the down-regulation of tubulin levels and disruption of microtubule structures caused by PrP mutants.

CK2 shows disease-associated alteration in the scrapie experimental rodents and human prion diseases. In this study, mammalian expressing plasmids for human CK2 subunits, CK2α and CK2ß, were generated. Immunoprecipitation assays revealed stronger signals of PrP-CK2α complexes in the HEK293 cells co-transfected with plasmids expressing CK2α and various PrP constructs, including PG5, CytoPrP, PG9, and PG12. Meanwhile, obviously weaker signals of PrP-CK2ß complexes were also observed in the cells co-expressing CK2ß and PrPs. Tubulin-specific Western blots and immunofluorescence assays revealed that similar as the observations in the presences of PrP-specific siRNA, the abnormal PrPs-induced reductions of tubulin and disruptions of microtubule structures were completely restored in the cells when co-expressing CK2α and CK2ß. Moreover, co-expressions of CK2α and PrPs induced phosphorylation on p53 at the position of serine 6 (p53-Ser6), although much weaker than that in the cells expressing CK2α and CK2ß, while expressions of either PrPs or CK2 subunits did not change the cellular p53 level or induce phosphorylation on p53 at Ser9. Our data here verify again the molecular interaction between CK2 and PrP. Co-presences of CK2 subunits restore the down-regulated tubulin and disrupted microtubule structures caused by expressions of the abnormal PrP proteins in HEK293 cells.

Molecular interaction of TPPP with PrP antagonized the CytoPrP-induced disruption of microtubule structures and cytotoxicity.

BACKGROUND: Tubulin polymerization promoting protein/p25 (TPPP/p25), known as a microtubule-associated protein (MAP), is a brain-specific unstructured protein with a physiological function of stabilizing cellular microtubular ultrastructures. Whether TPPP involves in the normal functions of PrP or the pathogenesis of prion disease remains unknown. Here, we proposed the data that TPPP formed molecular complex with PrP. We also investigated its influence on the aggregation of PrP and fibrillization of PrP106-126 in vitro, its antagonization against the disruption of microtubule structures and cytotoxicity of cytosolic PrP in cells, and its alternation in the brains of scrapie-infected experimental hamsters. METHODOLOGY/PRINCIPAL FINDINGS: Using pull-down and immunoprecipitation assays, distinct molecular interaction between TPPP and PrP were identified and the segment of TPPP spanning residues 100-219 and the segment of PrP spanning residues 106-126 were mapped as the regions responsible for protein interaction. Sedimentation experiments found that TPPP increased the aggregation of full-length recombinant PrP (PrP23-231) in vitro. Transmission electron microscopy and Thioflavin T (ThT) assays showed that TPPP enhanced fibril formation of synthetic peptide PrP106-126 in vitro. Expression of TPPP in the cultured cells did not obviously change the microtubule networks observed by a tubulin-specific immunofluorescent assay and cell growth features measured by CCK8 tests, but significantly antagonized the disruption of microtubule structures and rescued the cytotoxicity caused by the accumulation of cytosolic PrP (CytoPrP). Furthermore, Western blots identified that the levels of the endogenous TPPP in the brains of scrapie-infected experimental hamsters were significantly reduced. CONCLUSION/SIGNIFICANCE: Those data highlight TPPP may work as a protective factor for cells against the damage effects of the accumulation of abnormal forms of PrPs, besides its function as an agent for dynamic stabilization of microtubular ultrastructures.

Knockdown of prion protein (PrP) by RNA interference weakens the protective activity of wild-type PrP against copper ion and antagonizes the cytotoxicity of fCJD-associated PrP mutants in cultured cells.

Development of the pathogenesis of transmissible spongiform encephalopathies (TSEs) requires the presence of both the normal host prion protein (PrPC) and the abnormal pathological proteinase-K resistant isoform (PrPSc). Reduction of PrPC levels has been shown to extend survival time after prion infection. In this report, based on analysis of the known sequences of human PrP, we constructed two small interfering RNA (siRNA) duplexes targeting the segments of amino acids (aa) 108-114 (Ri2) and aa 171-177 (Ri3). Western blot analysis results revealed that these PrP-specific siRNAs could effectively knock down the levels of either endogenous PrP in human neuroblastoma SHSY-5Y cells or recombinant PrP transfected with the plasmid expressing the full-length human PrP in human embryonic kidney (HEK) 293T cells. Meanwhile, the two siRNAs also showed a significant effect on the reduction of the expression of the PrP-PG9 and PrP-PG12 familial Creutzfeldt-Jakob disease (CJD)-associated PrP mutants with four and seven extra octarepeats, in the cells transfected with the respective expression plasmids. MTT tests identified that knockdown of wild-type PrP by Ri2 and Ri3 did not change the cell growth capacities, but significantly decreased the cell resistances against the challenge of Cu2+. Co-expression of Ri2 and Ri3 partially antagonized the cytotoxicity caused by expressing PrP-PG9 and PrP-PG12 in the two cell lines. Moreover, the rescuing effectiveness of PrP siRNAs was time-related, with the more efficient antagonism of the cytotoxicity of fCJD-associated PrP mutants occurring at the early stages after transfection. The data shown here provide useful clues for seeking potential therapeutic tools for prion diseases.

[siRNA-mediated inhibition of ErbB2 expression and its effect on breast cancer cell growth].

AIM: To construct a vector of ErbB2 small interfering RNA(siRNA), and to investigate its effect on ErbB2 expression and the growth of ZR75-1 breast cancer cells. METHODS: Two ErbB2-siRNAs were designed and inserted into the pSliencer 2.1-U6 neo vector. After confirmed by restriction and DNA sequencing, siRNA vectors were co-transfected with the FLAG-ErbB2 expression vector into human embryonic kidney 293T cells, or transfected into SKBR3 and ZR75-1 breast cancer cells. The effects of siRNAs on the expression of ErbB2 were identified by Western blot and the proliferation of ZR75-1 cells was repressed. RESULTS: Two siRNAs could effectively inhibit the expression of exogenous and endogenous ErbB2 proteins, and could repress the growth of ZR75-1 cells. CONCLUSION: ErbB2 siRNAs effectively inhibit the expression of ErbB2, thus repressing the growth of ZR75-1 cells.

Heat shock protein 104 inhibited the fibrillization of prion peptide 106-126 and disassembled prion peptide 106-126 fibrils in vitro.

Amyloid-like fibrils have been associated with the pathogenesis of human prion diseases. Prion peptide of aa 106-126 (PrP106-126) exhibits many PrP(Sc)-like biochemical features, forming amyloid-like fibrils in vitro. Here, we found that the recombinant yeast-derived molecular chaperon Hsp104 inhibited significantly the fibril assembly of the synthetic PrP106-126 peptide by dynamic ThT assays in vitro. EM assays revealed almost no fibril-like structure after incubation of the synthetic PrP106-126 peptides with Hsp104 for 12h. Circular dichroism assays identified that treatment of Hsp104 shifted the secondary structure of PrP106-126 fibrils from ß-sheet to a random coil. MTT tests confirmed that interaction of PrP106-126 with Hsp104 maintained the toxicity of PrP106-126 on human neuroblastoma cell line SK-N-SH. Additionally, Hsp104 was able to disassemble the mature PrP106-126 fibrils in vitro, leading to recovering the cytotoxicity of PrP106-126 on SK-N-SH cells. Our study provides the molecular evidences that the yeast-derived Hsp104 can interfere in the fibril assembly and disassembly of human PrP106-126 segment.

[Establishment of estrogen receptor alpha trans-activation system].

OBJECTIVE: To construct estrogen receptor alpha (ERalpha) trans-activation system. METHODS: The full length ERalpha and its different function regions [(transcriptional activation function 1 (AF1), DNA binding domain (DBD), and transcriptional activation function 2 (AF2)] were amplified from pcDNA3-ERalpha by PCR and cloned into the pGAL vector. The expressions of the recombinant plasmids constructed were detected via immunoblotting. The 293T cells transfected with recombinant plasmids of full length ERalpha, its different function regions and empty vector were divided into 5 groups; each group was divided into 2 parts which were treated with or without estrogen (E(2)). The transcriptional activity of each group was detected in 293T cells after the recombinant plasmid was co-transfected with 0.2 microg of estrogen receptor element luciferase (ERE-LUC) and 0.1 microg of plasmid expressing beta-galactosidase and treated with or without 10 nmol/L E(2) for 24 hours. RESULTS: The full length ERalpha and its different function regions were expressed in the 293T cells. Compared with the empty pGAL vector, the transcription activities of full length ERalpha, AF1, AF2 and DBD recombinant plasmids were raised about 20.44 +/- 1.01, 2.09 +/- 0.11, 8.09 +/- 0.30 and 1.05 +/- 0.09 fold, respectively, with the induction of E(2) after transfection in the 293T cells. CONCLUSION: The trans-activation system of ERalpha has been successfully established.