Epidemiological studies have demonstrated strong associations between exposure to ambient fine particulate matter (PM2.5) and cardiac disease. To investigate the potential mechanism of cardiac fibrosis induced by PM2.5, we established PM2.5 exposure models in vivo and in vitro, and then cardiac fibrosis was evaluated. The ferroptosis and ferritinophagy was detected to characterize the effects of PM2.5 exposure. The results indicated that PM2.5 exposure could induce cardiac fibrosis in mice. YY1 was induced by PM2.5 exposure and then increased NCOA4, a cargo receptor for ferritinophagy, which interacted with FHC and promoted the transport of ferritin to the autophagosome for degradation. The release of large amounts of free iron from ferritinophagy led to lipid peroxidation directly via the Fenton reaction, thereby triggering ferroptosis. Moreover, siNCOA4 could partly restore the FHC protein level in HL-1 cells and inhibit the occurrence of downstream ferroptosis. Func-tionally, NCOA4 knockdown inhibited ferroptosis and alleviated HL-1 cell death induced by PM2.5. Ferroptosis inhibitor (Ferrostatin-1) could reverse the promoting effect of ferritinophagy mediated ferroptosis on cardiac fibrosis induced by PM2.5 exposure in mice. Our study indicated that PM2.5 induced cardiac fibrosis through YY1 regulating ferritinophagy-dependent ferroptosis.

Accumulating evidence has suggested that men exposed to air pollution are associated with decreased sperm quality, and seminal plasma plays a pivotal role in maintaining sperm viability. However, the role of seminal plasma in air pollution related sperm quality decline remain unestablished. In current study, we recruited 524 participants from couples who underwent in vitro fertilization treatment due to female factors at a fertility clinic in China from March to August 2020. Conventional sperm parameters, total antioxidant capacity (T-AOC), malondialdehyde (MDA) and testosterone were measured using semen samples. The six main air pollutants (PM2.5, PM10, NO2, SO2, CO, O3) during four key periods of sperm development (meiotic stage, spermiogenesis stage, epididymal stage and total sperm cycle period) were estimated using inverse distance weighting method. Multiple linear regression models were employed to investigate the exposure-outcome relationships. And we found that PM10 exposures were negatively related to sperm total motility and the exposures of PM2.5 and PM10 were inversely associated with sperm progressive motility during epididymal stage. Furthermore, PM2.5 and PM10 exposures were positively associated with seminal plasma MDA and PM10 was negatively related to seminal plasma T-AOC during epididymal stage. PM2.5, PM10 and CO exposures during total sperm cycle period might relate to increased seminal plasma testosterone. Mediation analysis indicated seminal plasma MDA and T-AOC partially mediated PM10 associated reduction of sperm motility during epididymal stage. Our study suggested MDA and T-AOC of seminal plasma played a role in air pollution associated decline of sperm motility.

Epidemiological studies have established an association between chronic exposure to PM2.5 and male infertility. However, the underlying mechanisms were not fully revealed. In this study, we established mice models exposed to PM2.5 for 16 weeks, and a significant decrease in sperm quality accompanied by an increase in testosterone levels were observed after PM2.5 exposure. Moreover, treatment with ferrostatin-1 (Fer-1), a specific ferroptosis inhibitor, effectively mitigated PM2.5-induced testicular dysfunction in mice. And lipid peroxidation and ferritin accumulation were found to be significantly increased in Leydig cells of testes with a PM2.5-dose dependent manner. Further investigations revealed that TM-3 cells, a mouse Leydig cell line, were prone to ferroptosis after PM2.5 exposure, and the cell viability was partly rescued after the intervention of Fer-1. Furthermore, our results supported that the ferroptosis of TM-3 cells was attributed to the upregulation of ferredoxin 1 (FDX1), which was the protein transferring electrons to cytochrome P450 family 11 subfamily A member 1 to aid lysing cholesterol to pregnenolone at initial of steroidogenesis. Mechanically, PM2.5-induced FDX1 upregulation resulted in cellular ROS elevation and ferrous iron overload, which together initiated an autoxidation process of polyunsaturated fatty acids in the cell membrane of Leydig cells until the accumulated lipid peroxides triggered ferroptotic cell death. Simultaneously, upregulation of FDX1 promoted steroidogenesis and let to an increased level of testosterone. In summary, our work suggested that FDX1, a mediator involving steroidogenesis, was a key regulator in PM2.5-induced Leydig cells ferroptosis.

Long-term inhalation of fine particulate matter (PM2.5) can cause serious effects on the respiratory system. It might be attributed to the fact that PM2.5 could directly enter and deposit in lung tissues. We established models of PM2.5 exposure in vivo and in vitro to explore the adverse effects of ambient PM2.5 on pulmonary and its potential pathogenic mechanisms. Our results showed that PM2.5 exposure promoted the deposition of ECM and the increased stiffness of the lungs, and then led to pulmonary fibrosis in time- and dose- dependent manners. Pulmonary function test showed restrictive ventilation function in mice after PM2.5 exposure. After PM2.5 exposure, ALKBH5 was recognized by TRIM11 and then degraded through the proteasome pathway. ALKBH5 deficiency (ALKBH5-/-) aggravated restrictive ventilatory disorder and promoted ECM deposition in lungs of mice induced by PM2.5. And the YAP1 signaling pathway was more activated in ALKBH5-/- than WT mice after PM2.5 exposure. In consequence, decreased ALKBH5 protein levels regulated miRNAs and then the miRNAs-targeted YAP1 signaling was activated to promote pulmonary fibrosis induced by PM2.5.

Exposure to PM2.5 increases blood pressure (BP) and cardiovascular morbidity and mortality. We conducted a randomized controlled panel study in Shijiazhuang, China among 55 healthy college students randomly assigned to either the control (CON) or SPORTS group with intervention of 2000 m jogging in 20 min for 3 times in 4 days, and 3-round health examinations from November 15, 2020 to December 6, 2020. We aimed to evaluate whether moderate physical activity (PA) protected BP health against PM2.5 exposure and explore potential mechanisms through myokines and inflammation. Individual PM2.5 exposure was calculated based on outdoor and indoor PM2.5 concentration monitoring data as well as time-activity diary of each subject. In the CON group, the exposure-response curve for SBP was linear with a threshold concentration of approximately 31 μg/m3, while an increment of SBP level was 4.38 mm Hg (95%CI: 0.17 mm Hg, 8.59 mm Hg) at lag03 for each 10-μg/m3 increase in PM2.5, using linear mixed-effect models. For inflammatory indicators, PM2.5 exposure was associated with significant increases in eosinophil counts and proportion in CON group, but decreases in MCP-1 and TNF-α in SPORTS group. Meanwhile, higher myokines including CLU and IL-6 were observed in SPORTS group compared to the CON group. Further mediation analyses revealed that eosinophil counts mediated the elevated BP in CON group, whereas MCP-1 and TNF-α were also crucial mediating cytokines for the SPORTS group, as well as CLU and IL-6 acted as mediators on BP and inflammation indicators in SPORTS group. This study suggests that moderate PA could counteract the elevated BP induced by PM2.5 exposure via myokines-suppressed inflammation pathways.

Objective To evaluate the pregnancy outcomes of progestin-primed ovarian stimulation (PPOS) protocol for patients with endometrioma underwent in vitro fertilization/intra-cytoplasmic sperm injection embryo transfer (IVF/ICSI-ET). Design Observational retrospective cohort study. Setting University affiliated reproductive center. Study Participants 605 infertile patients with endometrioma underwent IVF/ICSI-ET from January 2016 to March 2021 were included in this study. Methods Multivariable logistic regression analyses were conducted to determine the independent effect of controlled ovarian stimulation (COS) protocols on reproductive outcomes of first embryo transfer (ET) cycles. The live birth was primary outcome, the implantation rate, biochemical pregnancy, clinical pregnancy and ongoing pregnancy were secondary outcomes. Results Compared to PPOS protocol, the probability of implantation showed no significant difference with ultra-long gonadotrophin-releasing hormone agonist (GnRHa) protocol and gonadotrophin-releasing hormone antagonist (GnRHant) protocol (OR 1.7, 95% CI 0.9-3.1, OR 1.2, 95% CI 0.7-2.1, respectively). The PPOS protocol was correlated with a significantly lower biochemical pregnancy and clinical pregnancy than ultra-long GnRHa protocol in the multivariable logistic regression analysis (OR 2.3, 95% CI 1.1-4.9, OR 2.4, 95% CI 1.1-5.3, respectively). However, there was no significant difference in terms of biochemical pregnancy, clinical pregnancy and ongoing pregnancy between PPOS and GnRHant protocol (OR 1.4, 95% CI 0.7-2.7, OR 1.3, 95% CI 0.7-2.4, OR 1.1, 95% CI 0.6-2.3, respectively). In addition, compared to PPOS protocol, ultra-long GnRHa protocol and GnRHant protocol demonstrated no statistical difference in ongoing pregnancy (OR 2.0, 95% CI 0.9-4.5, OR 2.1, 95% CI 0.6-2.3, respectively). Notably, the ultra-long GnRHa protocol was associated with a significant higher probability of live birth than PPOS protocol both in crude analysis and multivariable logistic regression analysis (OR 2.6, 95% CI 1.3-5.1, OR 2.5, 95% CI 1.1-5.7, respectively). Nevertheless, no statistical difference was found in live birth between PPOS and GnRHant protocol either in crude analysis and multivariable logistic regression analysis (OR1.2, 95% CI 0.6-2.3, OR 1.2, 95% CI 0.6-2.5, respectively). Conclusions Based on the reproductive outcomes of the first ET cycles in patients with endometrioma, PPOS protocol may associated with inferior reproductive outcomes in terms of biochemical pregnancy, clinical pregnancy and live birth than ultra-long GnRHa protocol. However, there was no significant difference in implantation rate, clinical pregnancy, ongoing pregnancy and live birth between PPOS and GnRHant protocol.

Both long-term exposure to air pollution and abnormal fasting blood glucose (FBG) are linked to dyslipidemia prevalence. However, the joint role of air pollution and FBG on dyslipidemia remains unknown clearly. In this study, we aimed to test whether abnormal FBG could enhance the risks of long-term exposure to air pollutants on dyslipidemia in general Chinese adult population. The present study recruited 8917 participants from 4 cities in Hebei province, China. Participants' individual exposure to air pollutants was evaluated by the Empirical Bayesian Kriging statistical model in ArcGIS10.2 geographic information system. Dyslipidemia was defined according to Guidelines for the Prevention and Treatment of Dyslipidemia in Chinese Adults. Subjects were grouped into normal, prediabetes, diabetes according to FBG level. Generalized linear models were applied to analyze the interaction of air pollutants and FBG on dyslipidemia prevalence. The prevalence of dyslipidemia was 43.83% in our investigation. After adjusting all covariates, we found the risk of four air pollutants (PM2.5, PM10, NO2, SO2) on dyslipidemia prevalence was stronger as higher FBG level, and the adjusted odd ratio of interaction (ORinter (95% CI)) between PM2.5, PM10, NO2, SO2 and FBG levels on dyslipidemia was 1.171 (1.162, 1.189), 1.119 (1.111, 1.127), 1.124 (1.115, 1.130), 1.107 (1.098, 1.115), respectively. Stratified analyses indicated the modifying effects of FBG on the association of air pollution with dyslipidemia were stronger among male, less than 65 years old, overweight/obesity (all Pinter<0.1). Our study concluded that high FBG levels strengthened the risk of long-term exposure to air pollution on dyslipidemia, especially more noticeable in male, less than 65 years old, overweight.

Background: Congenital anomalies (CAs) are the leading causes for children's disabilities and mortalities worldwide. The associations between air pollution and CAs are not fully characterized in fetuses born by in vitro fertilization (IVF) who are at high risk of congenital anomalies. Methods: We conducted a cross-sectional study including 16,971 IVF cycles from three hospitals in Hebei Province, China, 2014-2019. Air quality data was obtained from 149 air monitoring stations. Individual average daily concentrations of PM2.5, PM10, NO2, SO2, CO, and O3 were estimated by spatiotemporal kriging method. Exposure windows were divided into 5: preantral follicle period, antral follicle period, germinal period, embryonic period and early fetal period. Logistic generalized estimating equations were used to estimate the associations between air pollutants and overall or organ-system specific congenital anomalies. Negative control exposure method was used to detect and reduce bias of estimation. Results: We found increasing levels of PM2.5 and PM10 were associated with higher risk of overall congenital anomalies during early fetal period, equating gestation 10-12 weeks (OR: 1.05, 95% CI: 1.02-1.09, p = 0.013 for a 10 mu g/m3 increase of PM2.5; OR: 1.03, 95% CI: 1.01-1.06, p = 0.021 for a 10 mu g/m3 increase of PM10). Cleft lip and cleft palate were associated with PM10 in germinal period and early fetal period. The CAs of eye, ear, face and neck were related to CO in preantral follicle stage. We did not find an association between chromosome abnormalities and air pollution exposure. Conclusions: We concluded that ambient air pollution was a risk factor for congenital anomalies in the fetuses conceived through IVF, especially exposure in early fetal period.

Background:Infertility is an important social problem. Asthenozoospermia (AZS) is a common pathological cause of male infertility, but its pathogenesis is unclear. Shenjing Guben Wan (SJGBW), a traditional Chinese medicine, has shown remarkable effects during the clinical treatment of oligozoospermia or AZS.Methods:In this study, clinical evaluations were carried out on 184 AZS patients receiving SJGBW treatment, including sperm count, sperm quality, and pregnancy rate. Also, ornidazole was used to build an AZS mouse model, and SJGBW treatment was administered. The sperm quantity and fertility of mice in different groups were evaluated; a cholecystokinin octapeptide-8 (CCK-8) experiment was carried out to test the activity of seminiferous epithelium Sertoli cells, and immunohistochemistry and the Terminal Deoxynucleotidyl Transferase-mediated dUTP Nick-End Labeling (TUNEL) method were employed to test the pathological information and expression of the Sertoli cell surface marker in the testicular tissues of mice in each group.Results:The sperm vitality, progressive sperm motility, and sperm morphology of patients who received SJGBW treatment were all improved (P<0.05). In the AZS group, the average sperm count, sperm vitality, pregnancy rate, and female mouse litters were all lower relative to mice in the control group. Following SJGBW treatment, the average sperm count, sperm vitality, pregnancy rate, and female mouse litters of mice in the AZS group were all significantly improved. The cytobiological experimental results showed that compared with the serum of normal male mice in the control group, the drug serum containing SJGBW could improve the cell vitality and proliferative ability of seminiferous epithelium Sertoli cells in AZS mice. Furthermore, the TUNEL results showed that the seminiferous tubule Sertoli cells and mesenchymal cells of the AZS mice exhibited the most significant apoptosis, which was alleviated following SJGBW treatment. Moreover, the levels of Sertoli cell marker, SOX9, and anti-apoptosis protein, Bcl2, in SJGBW-treated mice were both higher than that in AZS mice.Conclusions:SJGBW can promote the development and maturation of germ cells by facilitating the proliferation of Sertoli cells in AZS patients, thereby improving the fertility of these patients.

Although the association between ambient particulate matter and metabolic syndrome (MetS) has been investigated, the effect of particulate matter (PM) on MetS is inconclusive. We conducted a systematic review and meta-analysis to study the association between long-term ambient PM exposure and MetS risk. The data from five databases were extracted to analyze the association between ambient PM exposure and MetS risk. A random-effects model was performed to estimate the overall risk effect. The present systematic review and meta-analysis illustrated that an increase of 5 μg/m3 in annual PM2.5 or PM10 concentration was associated with 14% or 9% increases of MetS risk, respectively (PM2.5, RR = 1.14, 95%CI [1.03, 1.25]; PM10, RR = 1.09, 95%CI [1.00, 1.19]). The population-attributable risk (PAR) was 12.28% for PM2.5 exposure or 8.26% for PM10 exposure, respectively. There was statistical association between PM2.5 exposure and risk of MetS in male proportion ≥50%, Asia, related disease or medication non-adjustment subgroup as well as cohort study subgroups, respectively. The significant association between PM10 exposure and risk of MetS was observed in male proportion ≥50% and calories intake adjustment subgroups, respectively. Sensitivity analyses showed the robustness of our results. No publication bias was detected. In conclusion, there was positive association between long-term PM exposure and MetS risk. 12.28% of MetS risk could be attributable to PM2.5 exposure.

[背景]环境和饮食中多种金属的暴露与抑郁、焦虑的发生有关,但血清金属元素含量与抑郁、焦虑症状的发生以及神经递质水平的关系尚不明确.[目的]分析大学生血清金属元素含量与抑郁、焦虑量表评分及血清神经递质水平的关联.[方法] 2017年12月招募河北省某医科大学86名在读本科学生,填写一般情况调查问卷、抑郁自评量表(PHQ-9)和焦虑自评量表(GAD-7).采集空腹静脉血,离心分离血清,采用电感耦合等离子体-质谱法检测血清中11种金属元素含量,采用酶联免疫吸附测定法检测血清中皮质醇(CORT)、多巴胺(DA)和5-羟色胺(5-HT)的含量.采用套索(LASSO)模型筛选与PHQ-9、GAD-7评分及神经递质相关的金属元素,采用多元线性回归分析筛选出的血清金属元素含量与抑郁和焦虑量表评分、神经递质水平的关联.采用错误发现率(FDR)控制多重假设检验造成的假阳性率.[结果]本次调查中10.465％的大学生存在抑郁症状,5.814％的大学生存在焦虑症状.LASSO模型筛选出钴(Co)、砷(As)、硒(Se)、钼(Mo)和镉(Cd)含量与PHQ-9评分相关,As和铅(Pb)含量与GAD-7评分相关,Co和Cd含量分别与CORT和DA水平相关,Mo、Cd、锡(Sn)和锑(Sb)含量与5-HT水平相关.多元线性回归模型分析显示,校正年龄、性别、体重指数(BMI)后,Cd每增加10 μg·L-1,PHQ-9量表评分增加2.180 (95％CI:1.220～3.140)分;As每增加10μg·L-1,PHQ-9量表评分增加0.010 (95％ CI:0.001～0.020)分;Se每增加10μg·L-1,PHQ-9量表评分增加0.010 (95％CI:0.001～0.010)分;Co每增加10μg·L-1,PHQ-9量表评分增加0.930(95％ CI:0.410～1.460)分,CORT水平下降2.990 (95％CI:-5.690～-0.330) ng·L1;Mo每增加10μg·L-1,PHQ-9量表评分增加0.190 (95％ CI:0.030～0.340)分,5-HT水平升高0.980 (95％CI:0.230～1.730) ng·L-1 (PFDR＜0.05).[结论]部分大学生存在抑郁、焦虑表现.血清中金属元素Cd、Co、As、Se和Mo含量与PHQ-9评分存在关联,但未见血清中金属元素与GAD-7评分的关联;Co含量与CORT水平有负向关联,Mo含量与5-HT水平有正向关联.研究结果说明血清金属元素含量与大学生抑郁情绪有关.

Emerging evidence demonstrated that traffic-related air pollution induced adverse effects on cardiovascular system. We designed a population-based cross-sectional study to explore the association between residential proximity to major roadways, traffic density and the prevalence of valvular heart disease (VHD). A total of 34040 subjects from a Rural Health Project between 2013 and 2018 were collected. According to the inclusion and exclusion criteria, 4158 participants were enrolled in the final analysis. And we calculated the subjects’ proximity to major roadways and collected the traffic density on the major roadways. Transthoracic echocardiography (TTE) was performed to diagnose the VHD, according to the current AHA/ACC (the American Heart Association and the American College of Cardiology) guidelines. Differences between groups were examined by the one-way ANOVAs for continuous variables and the chi-square tests for categorical variables. A logistic regression models were used to assess the associations. The stratified analysis by age and sex were conducted to further analyze the association. The restricted cubic spline analysis was performed to further evaluate the association between road way distance and VHD. Bonferroni test was used to adjust the significance level. The subjects closer to the major roads had the higher risk of tricuspid regurgitation (TR) (odds risk, OR = 1.519, 95% confidence intervals, 95%CI: 1.058-2.181), especially in female. The risk of VHD was positive (high traffic density VS low traffic density, OR = 1.799, 95%CI: 1.221-2.651), especially in female. In addition, the high traffic density was associated with the risk of mitral regurgitation (MR) (OR = 1.758, 95%CI: 1.085-2.848). The restricted cubic spline analysis found a threshold distance of about 300 m, where had the lowest risk of VHD, aortic regurgitation (AR), MR, TR. Our results found a positive association between traffic-related air pollution and VHD especially in female.

Maternal exposure to air pollution is associated with poor reproductive outcomes in in vitro fertilization (IVF). However, the susceptible time windows are still not been known clearly. In the present study, we linked the air pollution data with the information of 9001 women receiving 10,467 transfer cycles from August 2014 to August 2019 in The Second Hospital of Hebei Medical University, Shijiazhuang City, China. Maternal exposure was presented as individual average daily concentrations of PM2.5, PM10, NO2, SO2, CO, and O3, which were predicted by spatiotemporal kriging model based on residential addresses. Exposure windows were divided to five periods according to the process of follicular and embryonic development in IVF. Generalized estimating equation model was used to evaluate adjusted odds ratios (ORs) and 95% confidence intervals (CIs) for association between clinical pregnancy and interquartile range increased average daily concentrations of pollutants during each exposure period. The increased PM2.5 (adjusted OR = 0.95, 95% CI: 0.90, 0.99), PM10 (adjusted OR = 0.93, 95% CI: 0.89, 0.98), NO2 (adjusted OR = 0.89, 95% CI: 0.85, 0.94), SO2 (OR = 0.94, 95% CI: 0.90, 0.98), CO (adjusted OR = 0.93, 95% CI: 0.89, 0.97) whereas decreased O3 (OR = 1.08, 95% CI: 1.02, 1.14) during the duration from preantral follicles to antral follicles were the strongest association with decreased probability of clinical pregnancy among the five periods. Especially, women aged 20-29 years old were more susceptible in preantral-antral follicle transition stage. Women aged 36-47 years old were more vulnerable during post-oocyte retrieve period. Our results suggested air pollution exposure during preantral-antral follicle transition stage was a note-worthy challenge to conceive among females receiving IVF.

Evidences on the association of air pollutants and semen quality were limited and mechanism-based biomarkers were sparse. We enrolled 423 men at a fertility clinic in Shijiazhuang, China to evaluate associations between air pollutants and semen quality parameters including the conventional ones, sperm mitochondrial DNA copy number (mtDNAcn), sperm telomere length (STL) and seminal spermatogenic cells. PM2.5, PM10, CO, SO2, NO2 and O3 exposure during lag0-90, lag0-9, lag10-14 and lag70-90 days were evaluated with ordinary Kringing model. The exposure-response correlations were analyzed with multiple linear regression models. CO, PM2.5 and PM10 were adversely associated with conventional semen parameters including sperm count, motility and morphology. Besides, CO was positively associated with seminal primary spermatocyte (lag70-90, 0.49; 0.14, 0.85) and mtDNAcn (lag0-90, 0.37; 0.12, 0.62, lag10-14, 0.31; 0.12, 0.49), negatively associated with STL (lag0-9, -0.30; -0.57, -0.03). PM2.5 was positively associated with mtDNAcn (0.50; 0.24, 0.75 and 0.38; 0.02, 0.75 for lag0-90 and lag70-90) while negatively associated with STL (lag70-90, -0.49; -0.96, -0.01). PM10 and NO2 were positively associated with mtDNAcn. Our findings indicate CO and PM might impair semen quality testicularly and post-testicularly while seminal spermatogenic cell, STL and mtDNAcn change indicate necessity for more attention on these mechanisms.

Objective: To analyze the pregnancy outcome of intracytoplasmic sperm injection(ICSI)with spermatozoa from different sources on pregnancy outcome.Methods:The data of 859 cycles treated by ICSI with spermatozoa from different sources were retrospectively were analyzed.The cycles were divided into 3groups:699cycles with ejaculated sperm group(group A),114 cycles with percutaneous epididymal spermatozoa aspiration group(group B),and 46 cycles with testicular spermatozoa aspiration group(group C).Results:The normal fertilization(2PN)rates in groups A(82.4%)was significantly higher than that in group B(76.2%)and group C(71.8%)(P0.05).The cleavage rates of group A(98.6%)was also higher than that of group B(97.6%)and group C(98.3%),and a significant difference was only found between group A and group B(P0.05).There was no significant difference in available embryo rate and good quality embryo rate among the three groups(P0.05).The implantation rate in group A,B,C was32.5%,37.7% and 41.7% respectively,and there was no significant difference among the three groups(P0.05).The pregnancy rate was not significantly different among group A(50.8%),group B(56.1%)and group C(60.9%)(P0.05).Conclusions: The spermatozoa from ejaculation,epididymis or testis for ICSI would affect the fertilization rate.The similar embryo rate,good quality embryo rate,and pregnancy outcomes could be obtained,when the spermatozoa from different sources are used for ICSI.