Objective To detect the level of cytokines and their receptors secreted by peripheral blood CD8 + T lymphocytes from vitiligo patients,and to evaluate the influence of tea polyphenols on their secretion.Methods CD8+ T lymphocytes were isolated from the peripheral blood of 12 patients with progressive vitiligo,12 patients with stable vitiligo and 10 healthy controls,and cultured in vitro for 20 days.Then,some CD8+ T lymphocytes were treated with tea polyphenols of 100 mg/L for two days.Enzyme-linked immunosorbent assay (ELISA) was carried out to determine the levels of tumor necrosis factor (TNF)-α,interferon (IFN)-γ and interleukin-2 receptor (IL-2R) in the culture supernatants of CD8+ T lymphocytes before and after the treatment with tea polyphenols.Analysis of variance and t test were conducted to assess differences in these parameters among these groups and changes between pretreatment and posttreatment,respectively.Results There was a sequential decrease in the level of TNF-α ((191.302 ± 6.077) vs.(175.966 ± 2.467) vs.(173.664 ± 3.600) ng/L,F =4.784,P ＜ 0.05),but a sequential increase in that of IFN-γ ((280.182 ± 36.070) vs.(371.670 ± 24.352) vs.(447.147 ± 8.432) ng/L,F=9.036,P＜ 0.01) and IL-2R ((8.375 ± 0.161) vs.(8.845 ± 0.161) vs.(9.345 ±0.125) ng/L,F =9.639,P ＜ 0.01) in the culture supernatant of CD8+ T lymphocytes from patients with progressive vitiligo to patients with stable vitiligo and healthy controls.After two days of tea polyphenol treatment,a statistical decrease was observed in the level of TNF-α in the culture supernatant of CD8 + T lymphocytes from patients with progressive vitiligo ((164.797 ± 1.784) ng/L,P ＜ 0.01),patients with stable vitiligo ((166.150 ± 3.576) ng/L,P ＜ 0.05) and healthy controls ((155.028 ± 5.759) ng/L,P ＜ 0.05),but no statistical changes were noted for IFN-γor IL-2R (all P ＞ 0.05) despite a slight elevation in the level of IFN-γin the culture supernatant of CD8+ T lymphocytes from patients with stable vitiligo and in that of IL-2R from all the three groups as well as a mild reduction in that of IFN-γ from patients with progressive vitiligo and healthy controls.Conclusions The changes of cytokines and their receptors secreted by CD8 + T lymphocytes may be associated with the induction and progression of vitiligo.Tea polyphenols may treat vitiligo via suppressing the secretion of TNF-α by CD8+ T lymphocytes.

Objective To evaluate the protective effects of tea polyphenols against the destruction of melanocytes by CD8+ T cells from vitiligo patients.Methods Skin tissue was resected from the margin of vitiligo lesions followed by the isolation and culture of CD8+ T lymphocytes,and from the normal skin of vitiligo patients followed by the isolation and culture of melanocytes.Flow cytometry was carried out to evaluate the purity of CD8+ T cells.The melanocytes were cocultured with the CD8 + T cells at different ratios followed by the evaluation of killing effect of CD8+ T cells.Various concentrations (200 and 400 μg/ml) of tea polyphenols were added into the co-culture system of CD8+ T cells and melanocytes at a ratio of 5 ∶ 1 followed by an additional culture of 48 hours.Then,flow cytometry was performed to detect the apoptosis in melanocytes in the coculture system.Results CD8+ T lymphocytes were successfully obtained from the marginal area of vitiligo lesions with a purity of more than 90％,which highly expressed the antigens CD137 and CD69.The coculture with CD8+ T cells markedly accelerated the apoptosis in melanocytes,while the accelerative effect was inhibited by tea polyphenols of 200 and 400 μg/ml.Conclusions The CD8+ T cells infiltrating the edge of vitiligo lesions display a potential destructive effect on autologous melanocytes from vitiligo patients,and tea polyphenols have a protective effect against the destruction of melanocytes by CD8+ T cells.

Objective To evaluate clinical value of denaturing high performance liquid chromatography (DHPLC) used in detecting transforming growth factor beta receptor 3 (TGFBR-3) exons 11 and 12 polymorphism in women with idiopathic premature ovarian failure (POF).Methods From Feb.2009 to Dec.2011,110 patients with idiopathic POF undergoing treatment at Shenzhen Maternal & Child Health Institute affiliated to Southern Medical University were enrolled as POF group in this study.In the mean time,110 women under 40 years old with normal hormonal level and menstrual cycles as control group.The exons 11 and 12 of TGFBR-3 gene polymorphism were screened by using DHPLC,and results of DNA sequencing was as golden standard.Some related indexes were calculated,such as sensitivity,specificity,false negative value,false positive value,Youden index,positive predictive value,and negative predictive value.At the same time,20％ of the tested specimens were chosen randomly and detected by DHPLC again.The value of Kappa index were calculated by comparing the results between the first and second DHPLC analysis.Results The exon 11 of TGFBR-3 were not identified gene polymorphism and two nucleotide polymorphisms were identified in exon 12.For 2022 T/C polymorphism,the frequencies of CC with 0.9％ (1/110),TC with 22.7％ (25/110),TT with 76.4％ (84/110),Cwith12.3％ (27/220) and T with 87.7％ (193/220) in POF group were significantly different from CC with 0,TC with 9.1％ (10/110)and TT with 90.9％ (100/110),C with 4.5％ (10/220) and T with 95.5％ (210/220) in control group (all P ＜ 0.05).Allelic and genotypic frequencies of 2161-75 C/T were not differed significantly between the two groups (all P ＞ 0.05).As DNA sequencing as golden standard,DHPLC showed that the sensitivity was 100％,specificity was 97.9％,Youden index was 97.9％,positive predictive value was 96.3％,negative predictive value was 100％,and Kappa index was 0.888 (P ＜ 0.05).Conclusion DHPLC analysis is higher validity,reliability and practicability method in detecting TGFBR-3 polymorphism in idiopathic premature ovarian failure.

Objective To evaluate the effect of calcipotriol on the proliferation of and cytokine secretion by melanocytes and perilesional CD8+ cytotoxic T lymphocytes (CTLs) from patients with vitiligo.Methods Melanocytes isolated from abdominal skin and CD8+ CTLs from perilesional skin of patients with vitiligo were subjected to successive culture in vitro.After several passages,the melanocytes and CD8+ CTLs were cultured alone or in combination with or without the presence of various concentrations of calcipotriol for 24 to 48 hours.MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfopheny)-2H-tetrazolium,inner salt) method was used to evaluate the proliferative activity of cells,enzyme-linked immunosorbent assay to determine the levels of interleukin (IL)-6,tumor necrosis factor (TNF)-α and interferon (IFN)-γ in the culture supematant of cells,flow cytometry to detect cell apoptosis.Some co-cultured melanocytes and CTLs were treated with calcipotriol of 10-8 mol/L and anti-IL-6 antibody of various concentrations (0,1,2,2.5,5,10 mg/L) for two days followed by enumeration of cells.The concentrations of 108 and 10-9 mol/L (calcipotriol) were chosen for relevant tests.Results There was a marked apoptosis in MCs after coculture with CD8+ CTLs.The 24-hour treatment with calcipotriol of 104 and 10-9 mol/L had no obvious effect on the proliferation of melanocytes cultured alone (both P ＞ 0.05),but accelerated the proliferation of melanocytes cocultured with CTLs (both P ＜0.05) as well as that of CD8+ CTLs cultured alone or in combination with melanocytes (all P ＜0.05).A statistical decrease was observed in IL-6,TNF-α and IFN-γlevels in the supernatant of cocultured melanocytes and CTLs compared with those in the supernatant of melanocytes and CTLs cultured alone,and calcipotriol of 10-9 mol/L intensified the decrease in supernatant IL-6 level (t =2.89,P ＜0.05),but no statistical changes were noted for the level of TNF-α or IFN-γin the supernatant of the coculture system after treatment with calcipotriol of 104 or 104 mol/L compared with that before treatment (both P ＞ 0.05).In the coculture system pretreated with calcipotriol of 10-8 mol/L,the number of CD8+ CTLs significantly decreased (t =3.15,P ＜0.05),whereas that of melanocytes significantly increased (t =3.53,P ＜0.05) after the treatment with anti-IL-6 antibody of 5 mg/L.Conclusions Perilesional CD8+ CTLs have a specific cytotoxic effect on melanocytes,and calcipotriol may inhibit the cytotoxic effect of CD8+ CTLs by suppressing the secretion of IL-6,which may partly explain the therapeutic mechanism of calcipotriol for vitiligo.

Objective To assess the influence of length of the alanine tract of forkhead box E1 (FOXE1) gene on genetic susceptibility to idiopathic premature ovarian failure (POF). Methods Totally 110 patients with idiopathic POF were recruited between February 2009 and December 2012 at the Affiliated Shenzhen City Maternity and Child Healthcare Hospital of Southern Medical University. Controls (n=110) were individuals with normal menstrual cycles, normal FSH concentrations. The polyalanine tract and flanking sequence of FOXE1 were screened using the multiplex ligation-dependent probe amplification (MLPA) technique and direct sequence technique. Results The most frequent of FOXE1 polyalanine stretch length was 14 residues in both groups. The length of FOXE1 polyalanine reported in this study varied from 12 to 16 alanines, and three variants of FOXE1-polyalanine length, containing 12, 14, or 16 alanine residues, and 5 different genotypes were identified. The most common genotypes were 14/14 homozygote, occurring with the frequency of 81.8% (90/110) in the POF group, while 96.4% (106/110) in control subjects, respectively. The incidence of 14/14 genotypes of FOXE1-polyalanine was significantly lower in patients with POF (χ2=119.730, P=0.001) in comparison to the controls. There were significantly higher frequencies of the 16/16 genotypes in cases with POF [10.0% (11/110) versus 0; χ2=3.403, P=0.001], as compared with the controls. The FOXE1 14 alanine allele was significantly less common in the POF patient group than the controls [84.5% (186/220) versus 98.2% (216/220); χ2=25.923, P=0.001]. The FOXE1 16 alanine allele was significantly more common in the POF patient group than the controls [12.7% (28/220) versus 1.8% (4/220); χ2=19.412, P=0.001]. Conclusions The polymorphism of the polyalanine tract of FOXE1 gene have a certain relevance for the genetic aetiology of idiopathic POF.

Aim To observe the effect of astragalosides on proliferation and collagen synthesis of HSC-T6 cells driven using schistosoma japonicum soluble egg antigen-activated macrophage conditioned medium(SEA-MCM).Methods SEA-MCM was prepared by injection of Schistosoma japonicum SEA via mice peritoneal.The proliferation and collagen synthesis of HSC-T6 cells stimulated with SEA-MCM were measured using MTT colorimetric assay and()~3H-proline incorporation respectively.Results The proliferation and collagen production of HSC-T6 cells were significantly promoted by SEA-MCM.They were significantly suppressed after being treated with AST(32.5、65、130 mg?L~(-1)) showing concentration-dependent effect.Conclusion:Astragalosides may inhibit HSC-T6 cells proliferation and collagen production that was driven by SEA activated MCM in vitro.

Objective: To analyze the distribution and associated factors of high-risk genotypes of HPV in cervical infection among women in Shenzhen. Methods: The information on sociodemographic characteristics and HPV genotypes of HPV-positive women who participated cervical screening test from January 2014 to December 2016 was downloaded from Shenzhen Maternity and Child Healthcare Management Information System. According to the pathogenicity, the high-risk HPV genotypes were divided into 15 types including HPV 16, 18, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 66 and 68; and there were 6 low-risk genotypes including HPV 6, 11, 42, 43, 44, and 81. Chi-square tests were applied to compare the proportions of high-risk HPV infection among women who had different sociodemographic characteristics. A non-conditional logistic regression model was used to analyze the associated factors for high-risk HPV infection. Results: In total, all HIV positives received HPV genotyping, with an average age of (38.08±9.38) years old. There were 9 979 (93.9%) high-risk and 645 (6.1%) low-risk HPV infections. The proportions of HPV infections for high-risk type in each year were 91.5%, 93.8%, and 95.6%, increasing with the screening years (χ²=54.79, P<0.001). Multivariate logistic regression analysis showed that compared with women younger than 25 years old, women in other age groups (at age 26 to 30 years, 31 to 35 years, 36 to 40 years, 41 to 45 years, and 50 years or older) had increased risks of high-risk HPV infection, with OR (95%CI) of 1.67 (1.20-2.31), 1.49 (1.09-2.03), 1.71 (1.23-2.37), 1.65 (1.19-2.31), and 1.84 (1.26-2.67), respectively; compared with the married, single women had a decreased risk of high-risk HPV infection (OR (95%CI): 0.71 (0.50-1.00)); women received HPV testing in 2015 and 2016 showed higher risk of high-risk HPV infection than those in 2014 (OR (95%CI): 1.43 (1.17-1.74) and 2.03 (1.68-2.46)). The 5 most common HPV genotypes were HPV52 (25.1%, 2 670 cases), followed by HPV16 (19.2%, 2 041 cases), HPV58 (13.3%, 1 413 cases), HPV18 (9.9%, 1 048 cases), and HPV51 (9.3%, 993 cases). Conclusion Age, marital status, and screening year were associated with high-risk HPV infections. Besides HPV16 and HPV18, the prevention and control on HPV infections for HPV52, HPV58, and HPV51 should be prioritized in Shenzhen area.

Objective To develop the methodology for using TLDs and films to measure absorbed dose and 2D dose distribution produced by the multi-leaf collimator (MLC) in intensity modulated radiotherapy (IMRT),in order to provide the guidance on dose quality audit in IMRT.Methods A total of 30 different-typed accelerators were selected from 27 hospitals in Jiangsu,Sichuan,Hubei and Henan provinces,including 17 Varian accelerators,10 Elektas and 3 Simens.The same batch of films and TLDs were put in a 2 cm-thick solid plate for fixation and then loaded in a 15 cm × 15 cm × 15 cm polysyrene solid phantom supplied by International Atomic Energy Agency(IAEA) in terms of 90 cm SSD,19 cm depth,10 cm × 10 cm field at different doses.The standard dose curves wcrc established for film and TLD,respectively.The irradiated film was measured and then sent to the External Audit Group (EAG) in China.The TLD-and film-absorbed doses were compared with TPS-calculated doses.The 2D dose distribution on the IRMT MLC field was measured using films.The homogeneous phanton of 30 cn × 30 cm was scanned by CT and the image was transferred to the TPS.The IMRT was implemented with 6 Gy fractionated irradiation by placing a 25 cm × 25 cm film on the phantom surface at 95 cm SSD and at 5 cm depth.The irradiated film was sent to the IAEA dosimetry laboratory for measurement and calculation.2D dose distribution verification was conducted in thc same way consistent with the procedure of international multi-radiotherapy center.The 3 mm/3％ passing rate was calculated for 2D dose distribution and compared with the film-measured and TPS calculated result.Results IAEA requires the relative deviation of TLD and film measured absorbed dose are with in ± 5％.The relative deviation of TLD-and filmmeasured to TPS-calculated absorbed dose was within the range of ±0.7％-± 8.5％ and within ±0.3％ ±7.8％ in Jiangsu,Sichuan,Hubei and Henan provinces,respectively.IAEA requires the 3 mm/3％ passing rate of film-measured 2D distribution to be 90％.The result of the present study were up to 94.0％.The verification result of 2D dose distribution were within 70.0％-99.9％ in Sichuan,Jiangsu,Hubei and Henan provinces.Conclusions The adsorbed dose and 2D distribution can be audited using TLDs and films for MLC in IRMT.The method is scientific and applicable,economical and convenient for development of dose quality audit for a wide range of IRMT.

BACKGROUND: The biological and molecular characteristics of spread through air spaces (STAS), a newly recognized invasive mode of lung cancer, remain controversial. The aim of this study was to investigate the clinicopathological features and molecular characteristics of STAS in patients with pulmonary adenocarcinoma. METHODS: A total of 694 resected invasive non-mucinous lung adenocarcinomas diagnosed by clinicopathology from July 2019 to March 2021 in the First Affiliated Hospital of Guangzhou Medical University were collected, and the relationship between STAS and clinicopathological factors was analyzed. The state of protein expression of anaplastic lymphoma kinase (ALK) was detected by immunohistochemical method. Epidermal growth factor receptor (EGFR) was detected by amplification refractory mutation system-polymerase chain reaction (ARMS-PCR). ROS proto-oncogene 1-receptor (ROS1) was detected by reverse transcription-PCR (RT-PCR). RESULTS: A total of 344 STAS positive cases and 350 STAS negative cases were collected. By univariate analysis, STAS positivity was statistically associated with tumor maximum diameter (P<0.001), pleural invasion (P<0.001), lymphovascular invasion (P<0.001), nerve invasion (P=0.013), lymph node metastasis (P<0.001), clinical stage (P<0.001) and histological type (P<0.001). There was a statistical correlation between STAS and ALK protein expression (P=0.001). Multivariate analysis showed that STAS positive was correlated with pleural invasion (P=0.001), vascular invasion (P<0.001), lymph node metastasis (P=0.005)and ALK protein expression (P=0.032). CONCLUSIONS STAS is associated with highly aggressive biological behavior of lung adenocarcinoma, suggesting a poor prognosis.
Humans ; Lung Neoplasms/pathology* ; Lymphatic Metastasis ; Protein-Tyrosine Kinases ; Prognosis ; Neoplasm Staging ; Neoplasm Recurrence, Local/pathology* ; Proto-Oncogene Proteins ; Adenocarcinoma of Lung/pathology* ; Neoplasm Invasiveness ; Retrospective Studies

Objective: To develop the methodology for using TLD and radiochromic film to measure the planned target volume (PTV) and organ at risk (OAR) doses and 2D dose distribution in IMRT, in order to provide technical guidance on the dose quality audit in IMRT at home. Methods: China has participated in the research project launched by the international multi-radiotherapy centre (IMRC). IMRT polystyrene phantom provided by IAEA was scanned by CT scanner and then the scanned images were transmitted to TPS to outline prescribed dose to PTV and to OAR. The former was limited to 400 cGy while the latter limited to 200 cGy. IMRT was implemented with the phantom irradiated using 6 MV X-ray. The irradiated TLDs and films were sent to IAEA dosimerty laboratory for measurement and calculation. Jiangsu, Sichuan, Hubei and Henan provinces were selected to engage with this study for their variety of accelerators and highly skilled physicists. The procedures used were the same as in the IMRC and the irradiated TLDs and films were required to send to external audit group for measurement and calculation. Results: According to IAEA requirement, the relative deviations of the TLD-measured and TPS planned doses are within ±7.0% for PTV and OAR. The China′s research results at the IMRC have shown that the relative deviation of TLD-measured and TPS-planned values for the upper and lower PTV were -0.2% and 0.8%, respectively, consistent with the IAEA requirement, and the values for upper and lower OAR were -0.6% and -1.0%, respectively, consistent with the requirement. As the results have shown in four provinces, the relative deviations of the TLD-measured and TPS-planned were within 0 to 10.6% for upper and lower PTV and -0.6% to 20.9% for upper and lower OAR. According to IAEA requirement, the passing rate should be greater than 90% for 3 mm /3% for 2D dose distribution. China′s result at the IMRC is 100%, being excellent. The four provinces′ results have shown that 2D dose distribution pass rate of 3 mm/3% was in the range of 45.0%-100.0%. Conclusions The uses of TLD in quality audit for PTV and OAR doses and the radiochromic film in 2D dose distribution pass rate in IMRT are characterized by scientific feasibility, strong operability, easy-to-mail and data realibility. They are can be applied to quality assurance and audit in medical institutions in the country to on a large scale.