Objective:To investigate the value of robot-assisted laparoscopic indocyanine green sentinel lymph node (SLN) tracing in treating endometrial carcinoma.Methods:Thirty-two patients with early-staging endometrial carcinoma were operated with laparoscopic comprehensive staging laparotomy from January 2019 to December 2021. At the same time, the SLN detection was performed by near-infrared fluorescence imaging tracer technology, in which the tracer was indocyanine green. Sixteen cases were injected with indocyanine green before laparoscopic surgery, and 16 cases were injected with indocyanine green before robot-assisted laparoscopic surgery. The operation index, postoperative complications, prognosis, and lymph node dissection were compared between the two groups.Results:(1) The mean age of patients in the robot group was (54.7±8.1) years old, and was (54.9±8.8) years old in the laparoscopic group. There were no significant difference between the two groups ( t=0.06, P=0.951). (2) Intraoperative blood loss [(131±40) vs (169±57) ml], hemoglobin difference before and after surgery [(11.2±5.4) vs (15.5±5.7) g/L], the length of stay after operation [(6.2±1.3) vs (8.6±1.4) days] between the robot group and the laparoscopic group were compared, and the differences were statistically significant (all P<0.05). (3) SLNs were detected in all 16 patients in the robotic group, and a total of 41 SLNs were detected. SLNs were detected in 15 of the 16 patients in the laparoscopy group, and a total of 40 SLNs were detected. Compared with the laparoscopic group (15/16), the total detection rate of SLN in the robotic group (16/16), there were no statistical significance ( χ2=1.03, P=0.310). Compared with the laparoscopic group (7/15), the SLN bilateral detection rate in the robotic group (10/16), there were also no significant difference ( χ2=0.78, P=0.376). The number of lymph nodes detected in surgery group (16.6±4.1) were lower than those in the laparoscopy surgery group (21.0±7.1), while there were no statistically difference between the two groups ( χ2=2.01, P=0.054). There was no tumor metastasis in the resected lymph nodes and SLN between the two groups. The false negative rate of SLN in diagnosing endometrial cancer postoperative lymph node metastasis was 0, and the negative predictive value was 100%. (4) The pelvic and retroperitoneal lymph nodes were divided into five regions, which were the left pelvis, the right pelvis, the presacral region, the deep inguinal region, and the abdominal aorta. The numbers of SLN of unilateral detection and bilateral pelvic detection between two groups showed no significant differences (all P>0.05). The left pelvis had the most SLN imaging in both groups, followed by the right pelvis, para-aortic, and deep groin. (5) There was one patient in both robotic group and laparoscopic group with postoperative complications, which were urinary retention and pelvic lymph node cyst respectively. There were no significant differences in the incidence of complications between the two groups ( χ2=0.97, P=1.000). The median follow-up time after operation was 14 months (range 6-24 months). During the follow-up period, no local recurrence or distant metastasis was found between the two groups of endometrial cancer patients. Conclusions:Compared with the laparoscopic group, the robot group has less intraoperative blood loss and shorter postoperative hospital stay. The bilateral detection rate of SLN in the group was better than that of laparoscopy.

BACKGROUND:Piezo1,a mechanosensitive protein,is tightly connected to osteogenic differentiation,and it has been demonstrated that TAZ has a role in regulating osteogenic differentiation.It is unclear whether TAZ participates in the regulation of osteogenic differentiation of human bone marrow mesenchymal stem cells by Piezo1,so it is crucial to investigate its unique mechanism to prevent osteonecrosis of the femoral head. OBJECTIVE:To elucidate what function Piezo1 plays in osteogenic differentiation and TAZ expression in human bone marrow mesenchymal stem cells. METHODS:The siRNA targeting Piezo1 was constructed and transfected into 293T cells.The silencing efficiency was detected by RT-qPCR.The selected Piezo1-Home-2337 was packaged according to the silencing efficiency,and its optimal multiplicity of infection value was assayed by immunofluorescence staining.The packaged Piezo1 silencing recombinant lentivirus was transfected into human bone marrow mesenchymal stem cells,and its silencing effect was detected by RT-qPCR and western blot assay.Alizarin red staining,alkaline phosphatase activity analysis,immunofluorescence staining,RT-qPCR and western blot assay were utilized to analyze the effect of silencing Piezo1 on the osteogenic differentiation of human bone marrow mesenchymal stem cells. RESULTS AND CONCLUSION:(1)The mRNA and protein levels of Piezo1 in human bone marrow mesenchymal stem cells transfected by si-Piezo1 were decreased significantly,with a statistically significant difference compared with normal and negative control groups.(2)The alkaline phosphatase activity in the si-Piezo1 group was much lower and the calcium deposition in the si-Piezo1 group was significantly reduced compared with the negative control group.(3)The mRNA levels of osteogenesis-related genes including Runt-related transcription factor 2(Runx2),osteopontin(OPN),distal-less homeobox 5(DLX5),osteocalcin,β-catenin and Tafazzin(TAZ)in the si-Piezo1 group were significantly decreased compared with the negative control group.Afterward,the expression levels of TAZ and β-catenin protein in the si-Piezo1 group were down-regulated significantly compared with the negative control group,whereas the expression levels of p-TAZ and p-β-catenin protein in the si-Piezo1 group had the opposite condition.(4)The results of immunofluorescence staining showed that the expression of TAZ and β-catenin in human bone marrow mesenchymal stem cells in the si-Piezo1 group was less compared with the negative control group.(5)These findings indicate that Piezo1 can promote the osteogenic differentiation of human bone marrow mesenchymal stem cells.The osteogenic ability of human bone marrow mesenchymal stem cells is significantly reduced after silencing Piezo1,and the expression of TAZ is also reduced.

Objective: To establish a triple-color pseudovirion-based neutralization assay (PBNA) and evaluate its capability of detecting immunogenicity of the sera generated by the immunization of HPV 9-valent vaccine. Methods: HPV pseudovirus (PsVs) 6/11/16/18/31/33/45/52/58 with the encapsidated fluorescence expressing red fluorescent plasmid N31-MCHREEY, green fluorescent N31-EGFP or blue fluorescent N31-mTagBFP were generated. The concentration of HPV PsVs and the infection titers of HPV PsVs were detected by double-antibody sandwich ELISA and TCID₅₀, respectively. The single- and triple color HPV 16/33/45 PsVs were used to detect the neutralization titers of mice sera immunized with HPV 9-valent vaccine and confirmed the accuracy and specificity of the triple-color PBNAs. Then, the single- and triple color HPV 6/11/18/31/33/45/52/58 PsVs were employed to detect the neutralization titers of cynomolgus macaques sera immunized with HPV 9-valent vaccine and determined whether the triple-color PBNAs could be applied to evaluate the immunogenicity of the sera generated by the immunization of HPV9-valent vaccine. Results: The concentration of HPV16 PsVs encapsulating green, red or blue fluorescent plasmid was 5.0 to 6.0 μg/ml and HPV6/11/18/31/33/45/52/59 triple-color HPV PsVs was about 1.0 to 3.0 μg/ml. 9 types HPV PsVs containing EGFP, Mcherry or mTagBFP reporter plasmid were obtained and the concentration can meet the need of neutralization detection. 9 types single-color fluorescent HPV PsVs had similar infectivity against 293FT cells with the infection titer values between 1×10⁴ and 1×10⁵. The results of PBNAs showed that there was no significant difference in the anti-HPV neutralization titers of mice sera induced by HPV 9-valent vaccine between single-color and triple-color HPV16/33/45 PsVs (P>0.05). Similarly, there was also no significant difference in the anti-HPV neutralization titers of cynomolgus macaques sera induced by HPV 9-valent vaccine between single-color and triple-color HPV6/11/18/31/33/45/52/58 PsVs (P>0.05). Conclusion We successfully established the triple-color PBNAs and verified the accuracy and specificity of triple-color PBNAs consistent with single-color PBNAs. The triple-color PBNAs can be applied to evaluate the immunogenicity of HPV 9-valent vaccine's immune serum.