Cytospora is a genus including important phytopathogens causing severe dieback and canker diseases distributed worldwide with a wide host range. However, identification of Cytospora species is difficult since the currently available DNA sequence data are insufficient. Aside the limited availability of ex-type sequence data, most of the genetic work is only based on the ITS region DNA marker which lacks the resolution to delineate to the species level in Cytospora. In this study, three fresh strains were isolated from the symptomatic branches of Elaeagnus angustifolia in Xinjiang Uygur Autonomous Region, China. Morphological observation and multi-locus phylogenetic analyses (ITS, LSU, ACT and RPB2) support these specimens are best accommodated as a distinct novel species of Cytospora. Cytospora elaeagnicola sp. nov. is introduced, having discoid, nearly flat, pycnidial conidiomata with hyaline, allantoid conidia, and differs from its relatives genetically and by host association.

To establish the amplification-refractory mutation system quantitative real-time PCR (ARMS-qPCR) method based on qPCR technique for detecting the c.2T>C mutation of SLC25A13 gene and validate its diagnostic performance. According to the principle of ARMS-qPCR primer design, the specific primers were designed for the conserved sequence of SLC25A13. The c.2T>C mutation ARMS-qPCR detection assay of SLC25A13 gene and the corresponding Sanger sequencing system were established through the use of the synthetic plasmids of homozygous mutation and 200 human peripheral blood specimens which were verified by Sanger sequencing as templates, and the diagnostic efficacy of the qPCR assay was validated by using nucleic acid extracted from another 200 human peripheral blood specimens and the results obtained were compared with the Sanger sequencing results as the gold standard, and the consistency of the two detection methods was analyzed. The results showed that the qPCR assay could accurately identify artificial plasmids carrying different mutations of SLC25A13 gene, and distinguish between wild type SLC25A13 gene and the c.2T>C mutation. This method was used to detect the mutation status of SLC25A13 c.2T>C in human peripheral blood, and the detection results were 100% consistent with the Sanger sequencing results. Among the 200 blood samples, 8 samples (4%) carried the c.2T>C mutation of SLC25A13 gene and 192 samples (96%) did not carry it. In conclusion, the ARMS-qPCR test established in this study can quickly, simply and accurately detect the c.2T>C mutation of SLC25A13 gene, which is helpful for the diagnosis of citrin deficiency (CD).

To establish the amplification-refractory mutation system quantitative real-time PCR (ARMS-qPCR) method based on qPCR technique for detecting the c.2T>C mutation of SLC25A13 gene and validate its diagnostic performance. According to the principle of ARMS-qPCR primer design, the specific primers were designed for the conserved sequence of SLC25A13. The c.2T>C mutation ARMS-qPCR detection assay of SLC25A13 gene and the corresponding Sanger sequencing system were established through the use of the synthetic plasmids of homozygous mutation and 200 human peripheral blood specimens which were verified by Sanger sequencing as templates, and the diagnostic efficacy of the qPCR assay was validated by using nucleic acid extracted from another 200 human peripheral blood specimens and the results obtained were compared with the Sanger sequencing results as the gold standard, and the consistency of the two detection methods was analyzed. The results showed that the qPCR assay could accurately identify artificial plasmids carrying different mutations of SLC25A13 gene, and distinguish between wild type SLC25A13 gene and the c.2T>C mutation. This method was used to detect the mutation status of SLC25A13 c.2T>C in human peripheral blood, and the detection results were 100% consistent with the Sanger sequencing results. Among the 200 blood samples, 8 samples (4%) carried the c.2T>C mutation of SLC25A13 gene and 192 samples (96%) did not carry it. In conclusion, the ARMS-qPCR test established in this study can quickly, simply and accurately detect the c.2T>C mutation of SLC25A13 gene, which is helpful for the diagnosis of citrin deficiency (CD).

Objective:To investigate the effect of vascularized lymph node transfer (VLNT) combined with lymphatico-venous anastomosis (LVA) in the treatment of lymphedema after breast cancer surgery.Methods:The data of patients with upper limb lymphedema after breast cancer surgery who were treated in the Department of Reconstructive and Reconstructive Microsurgery, Ruikang Hospital Affiliated to Guangxi University of Traditional Chinese Medicine from July 2021 to July 2022 were retrospectively analyzed. According to different treatment methods, the patients were divided into LVA group and VLNT combined LVA group. Indocyanine green (ICG) near-infrared lymphography was performed on all affected limbs before surgery. In the LVA group, according to the results of ICG lymphography, 4 to 5 levels of the affected limb were selected, Z shaped incisions were made and dissescted until the subcutaneous fat layer. End-to-end or end-to-side anastomosis was performed between lymphatic vessels and subcutaneous venules under the microscope. In the VLNT combined LVA group, the branches of brachial artery and vein in the axillary region were marked. The inguinal flap with the superficial iliac circumflex vessel pedicle and 4-5 lymph nodes was dissected. End-to-end anastomoses of the superficial iliac circumflex vessel pedicle with the branches of brachial artery and vein were performed in the axillary region of the affected limb. LVA was performed according to ICG lymphography, the same as in the LVA group. The skin and soft tissue condition of the affected limb and the blood supply of the flap in the VLNT combined LVA group were observed after operation. The circumference of the upper arm (from the wrist to 32 cm above the wrist, every 4 cm, a total of 9 levels of circumference) and upper limb volume were measured before and after operation. SPSS 24.0 was used for data processing and analysis. Measurement data were expressed Mean±SD. The data before and after operation in the same group were compared by paired samples t test, and the comparison between the two groups was conducted by independent samples t test. P<0.05 was considered statistically significant. Results:A total of 14 female patients were enrolled, with 7 patients in each group. All cases were unilateral lymphedema. There were no significant differences in age, stage of disease, limb circumference and limb volume between the two groups before operation ( P>0.05). After operation, the skin and soft tissue condition of the affected limbs were good, and no complications such as erysipelas, cellulitis, or lymphangitis occurred. All flaps in the VLNT combined LVA group survived successfully, and the operation wounds healed well. There were no complications such as infection and necrosis of the flaps. One year after operation, the circumference and volume of the affected limb in the two groups were improved to varying degrees, and the circumference of the affected limb (wrist, upper wrist 4, 8, 12, 16, 20, 24, 28, 32 cm levels) in the VLNT combined LVA group was significantly smaller than that before operation ( P<0.01). In the LVA group, the circumference of the affected limb (wrist, upper wrist 4, 8, 12, 16, 20 cm levels) after operation was significantly smaller than that before operation ( P<0.05). The limb volumes of both groups were significantly reduced ( P<0.05). The comparison between the two groups showed that the reduction degree of postoperative affected limb cricumference (at the levels of 24, 28 and 32 cm above the wrist) and volume in the VLNT combined LVA group were more significant than those in the LVA group ( P<0.05). Conclusion:Compared with LVA alone, VLNT combined with LVA is more effective in the treatment of patients with upper limb lymphedema after breast cancer surgery.

Objective:To investigate the effect of vascularized lymph node transfer (VLNT) combined with lymphatico-venous anastomosis (LVA) in the treatment of lymphedema after breast cancer surgery.Methods:The data of patients with upper limb lymphedema after breast cancer surgery who were treated in the Department of Reconstructive and Reconstructive Microsurgery, Ruikang Hospital Affiliated to Guangxi University of Traditional Chinese Medicine from July 2021 to July 2022 were retrospectively analyzed. According to different treatment methods, the patients were divided into LVA group and VLNT combined LVA group. Indocyanine green (ICG) near-infrared lymphography was performed on all affected limbs before surgery. In the LVA group, according to the results of ICG lymphography, 4 to 5 levels of the affected limb were selected, Z shaped incisions were made and dissescted until the subcutaneous fat layer. End-to-end or end-to-side anastomosis was performed between lymphatic vessels and subcutaneous venules under the microscope. In the VLNT combined LVA group, the branches of brachial artery and vein in the axillary region were marked. The inguinal flap with the superficial iliac circumflex vessel pedicle and 4-5 lymph nodes was dissected. End-to-end anastomoses of the superficial iliac circumflex vessel pedicle with the branches of brachial artery and vein were performed in the axillary region of the affected limb. LVA was performed according to ICG lymphography, the same as in the LVA group. The skin and soft tissue condition of the affected limb and the blood supply of the flap in the VLNT combined LVA group were observed after operation. The circumference of the upper arm (from the wrist to 32 cm above the wrist, every 4 cm, a total of 9 levels of circumference) and upper limb volume were measured before and after operation. SPSS 24.0 was used for data processing and analysis. Measurement data were expressed Mean±SD. The data before and after operation in the same group were compared by paired samples t test, and the comparison between the two groups was conducted by independent samples t test. P<0.05 was considered statistically significant. Results:A total of 14 female patients were enrolled, with 7 patients in each group. All cases were unilateral lymphedema. There were no significant differences in age, stage of disease, limb circumference and limb volume between the two groups before operation ( P>0.05). After operation, the skin and soft tissue condition of the affected limbs were good, and no complications such as erysipelas, cellulitis, or lymphangitis occurred. All flaps in the VLNT combined LVA group survived successfully, and the operation wounds healed well. There were no complications such as infection and necrosis of the flaps. One year after operation, the circumference and volume of the affected limb in the two groups were improved to varying degrees, and the circumference of the affected limb (wrist, upper wrist 4, 8, 12, 16, 20, 24, 28, 32 cm levels) in the VLNT combined LVA group was significantly smaller than that before operation ( P<0.01). In the LVA group, the circumference of the affected limb (wrist, upper wrist 4, 8, 12, 16, 20 cm levels) after operation was significantly smaller than that before operation ( P<0.05). The limb volumes of both groups were significantly reduced ( P<0.05). The comparison between the two groups showed that the reduction degree of postoperative affected limb cricumference (at the levels of 24, 28 and 32 cm above the wrist) and volume in the VLNT combined LVA group were more significant than those in the LVA group ( P<0.05). Conclusion:Compared with LVA alone, VLNT combined with LVA is more effective in the treatment of patients with upper limb lymphedema after breast cancer surgery.

OBJECTIVE: To screen for small molecular compounds with selective inhibitory activity against cutaneous melanoma cells with BAP1 deletion. METHODS: Cutaneous melanoma cells expressing wild-type BAP1 were selected to construct a BAP1 knockout cell model using CRISPR-Cas9 system, and small molecules with selective inhibitory activity against BAP1 knockout cells were screened from a compound library using MTT assay. Rescue experiment was carried out to determine whether the sensitivity of BAP1 knockout cells to the candidate compounds was directly related to BAP1 deletion. The effects of the candidate compounds on cell cycle and apoptosis were detected with flow cytometry, and the protein expressions in the cells were analyzed with Western blotting. RESULTS: The p53 activator RITA from the compound library was shown to selectively inhibit the viability of BAP1 knockout cells. Overexpression of wild-type BAP1 reversed the sensitivity of BAP1 knockout cells to RITA, while overexpression of the mutant BAP1 (C91S) with inactivated ubiquitinase did not produce any rescue effect. Compared with the control cells expressing wild-type BAP1, BAP1 knockout cells were more sensitive to RITA-induced cell cycle arrest and apoptosis (P < 0.0001) and showed an increased expression of p53 protein, which was further increased by RITA treatment (P < 0.0001). CONCLUSION Loss of BAP1 results in the sensitivity of cutaneous melanoma cells to p53 activator RITA. In melanoma cells, the activity of ubiquitinase in BAP1 is directly related to their sensitivity to RITA. An increased expression of p53 protein induced by BAP1 knockout is probably a key reason for RITA sensitivity of melanoma cells, suggesting the potential of RITA as a targeted therapeutic agent for cutaneous melanoma carrying BAP1-inactivating mutations.
Humans ; Melanoma ; Skin Neoplasms ; Tumor Suppressor Protein p53 ; Apoptosis ; Cell Division ; Tumor Suppressor Proteins/genetics* ; Ubiquitin Thiolesterase/genetics*