{"paper_id":"3d0f1b22-49f7-472c-ab06-3282c806f358","body_text":"The demand for infertility treatment has grown exponentially in Brazil. Intrauterine\ninsemination (IUI) is a low complexity fertility treatment that is much less\ninvasive and inexpensive when compared to in vitro fertilization (IVF) ( Isa  et al ., 2014a ). Pregnancy\nrates after IUI vary widely due to multiple patient-related factors ( Isa  et al ., 2014a ;  2014b ;  Allen\n et al ., 1985 ;  Asante\n et al ., 2013 ;  Nuojua-Huttunen  et al ., 1999 ;  Schuffner  et al ., 2009 ). IUI can be indicated\nfor patients with different causes of infertility, including cervical factor\ninfertility, ovulatory dysfunction, endometriosis, infertility for immunological\ncauses, and idiopathic infertility ( Duran  et\nal ., 2003 ).\nSome factors may have a decisive role in the outcome of IUI, such as age, ovarian\nreserve, endometrial thickness, types and doses of gonadotropin, and sperm quality.\nAlthough several studies have searched for markers of success in IUI, age remains as\nthe best parameter to assess ovarian function and consequently the response to\nassisted reproductive technologies ( Deatsman\n et al ., 2016 ). Although some authors have linked\nhigher levels of AMH to higher pregnancy rates ( Bakas\n et al ., 2015 ), the predictive value of AMH remains\nquestionable in scientific literature ( Tremellen\n& Kolo, 2010 ).\nEndometrial thickness has been associated with higher embryo implantation and IUI\nsuccess rates. There is no consensus in the literature over an ideal cutoff value,\nalthough most studies agree on endometrial thickness greater than 7 mm ( Biswas  et al ., 2016 ;  Caetano  et al ., 2005 ).\nIUI without controlled ovarian stimulation is not recommended, as natural cycles\noffer no clinical advantage. Fertility rates are higher in stimulated cycles ( Rashidi  et al ., 2013 ;  Fritz & Speroff, 2011 ). In several\ncountries, it is mandatory to register IUI procedures. In Denmark, for example, IUI\ntreatments have been registered since 2007 ( Malchau\n et al ., 2014 ). REDLARA, the Latin American Network\nof Assisted Reproduction, reported 6,250 cycles of IUI in 2013 with a birth rate per\ncycle of 14.9% ( Zegers-Hochschild  et\nal ., 2016 ). In Brazil, outcomes and indications of IUI are\nscarce.\nThe aim of the study was to identify which parameters positively affected the\nclinical pregnancy rates of IUI cycles in a private assisted reproduction center and\nevaluate which couples should opt for IUI.\n\nThis retrospective observational study included 261 patients submitted to 381 IUI\ncycles with fresh or cryopreserved partner semen (IU-H) from January 2012 to\nFebruary 2017 in a private center in Curitiba-PR, Brazil. Heterologous\nintrauterine insemination procedures were not included, so as not to affect\npregnancy rates.\nThe local institutional review board approved the study. Patients were not\nrequired to give consent due to the retrospective nature of the study.\nThe included subjects were split into groups based on the cause of infertility,\nage, endometrial thickness (measured two days before IUI), and\nanti-Müllerian hormone (AMH) levels. AMH levels were measured in 125\npatients, and test results were not considered for IUI. The groups were further\ndivided between fresh  vs.  frozen semen and levels of sperm\nconcentration and motility.\nControlled ovarian stimulation was achieved by three different protocols: 1)\nInjectables, 2) Oral medication + injectables, 3) Oral medication. Group 1 was\ndivided into three subgroups: A) human menopausal gonadotropin (hMG) -\nMenopur ® , B) recombinant follicle stimulating hormone\n(rFSH) - Gonal ®  or Pergoveris ® , and C)\nurinary follicle stimulating hormone (uFSH) Fostimon ® . Group 2\ntook clomiphene citrate (CC) followed by the injectable medications cited above.\nGroup 3 took CC or Letrozole ® . The final outcome considered\nfor all variables was clinical pregnancy.\nSerial transvaginal ultrasound examination was performed on the second or third\nday of the cycle and on the sixth day after the start of controlled ovarian\nstimulation. The remaining tests varied according to the response each patient\nhad to the stimulation protocol. hCG (Choriomon ®  or\nOvidrel ® ) was administered when the mean diameter of the\ndominant follicle reached at least 18mm. IUI was scheduled 36h to 40h\nthereafter. Endometrial thickness was evaluated 48h before IUI, without later\nanalysis. Supplementation of the luteal phase with Utrogestan ® \nor Duphaston ®  was also performed.\nSemen samples were processed by density gradient centrifugation or sperm wash\naccording to the 2010 guidelines of the World Health Organization (WHO) for\nsemen processing ( WHO, 2010 ). Sperm wash\nwas performed in samples meeting the following criteria: increased viscosity,\nconcentration <15 million per ml, and motility <32%. Other samples\nunderwent density gradient centrifugation. In both procedures, the volume was\nmeasured after collection and concentration and motility were evaluated on\nMakler chambers with the aid of a light microscope.\nAfter complete liquefaction in density gradient centrifugation, the semen samples\nwere processed with density gradient medium (Isolate, Irvine Scientific,\nCalifornia) and sperm washing medium (Modified HTF Medium HEPES with Gentamicin,\nIrvine Scientific, California). In the processing technique, 90% and 45%\ncolloidal gradient were used. One mL of the lower phase gradient was transferred\ninto a sterile disposable conical-bottom centrifuge tube using a Pasteur\npipette. A second 1 mL layer of the upper phase was gently placed on top of the\nlower phase. Liquefied semen was gently placed onto the upper phase. The sample\nwas centrifuged for 20 minutes at 516G. The supernatant was discarded, and the\npellet was resuspended with a Pasteur pipette in 2.5 mL of HTF. Sperm wash was\nperformed with centrifugation for eight minutes at 516G. Then the supernatant\nwas discarded and the pellet resuspended with a Pasteur pipette in 0.5 mL of\nHTF. Sperm parameters were then evaluated according to the WHO criteria for\nIUI.\nData were organized on Microsoft Office Excel 2007 ®  and\nanalyzed on SPSS Statistics 22.0 ® . The results were expressed\nas frequencies and proportions for categorical and qualitative variables, and as\nmeasures of dispersion and central tendency for quantitative variables.\nThe Mann-Whitney test for quantitative variables was used to assess the\nrelationship between clinical/biological variables and pregnancy success. The\nnon-parametric Shapiro-Wilk test was chosen to test the normality of\nquantitative variables. The Chi-square or Fisher's exact test was used to test\nthe association between qualitative variables and the outcome of treatment.\nStatistical significance was attributed to comparisons with a\n p -value <0.05.\nLinear logistic regression models were used to determine the relationship between\noutcomes and the clinical/biological variables. Explanatory variables with a\nsignificance level of up to 20% in univariate analysis were included in the\nlogistic regression model. The results were expressed as odds ratios (OR).\nStatistical power analysis was performed to verify the ability of tests to reject\na false null hypothesis. Software program GPower 3.1 was used in the analysis of\nstatistical power from univariate and multivariate analysis.\n\nIdiopathic infertility was identified in 35.9% of the cases, followed by ovulatory\ndysfunction with 26.2% and endometriosis with 15.2%. Other less frequent causes were\ncervical factor, tubal factor, male factor and multiple factor infertility. There\nwas no statistical difference regarding the cause of infertility and the number of\npregnancies ( p =0.8). Fifty-six (14.6%) of the 381 cycles analyzed\nresulted in clinical pregnancy.\nPatients younger than 40 years accounted for 87.9% (335) of the sample and achieved\n54 clinical pregnancies (16.1%). The clinical pregnancy rate was five times higher\nin younger patients (<40 years) compared to their older peers (>40 years)\n( p =0.043). Among the patients with positive clinical\npregnancies, only two were older than 40 years ( Table 1 ). When the patients were split by age (≤29, 30-34, 35-39,\nand ≥40 years), most pregnancies - 26 (6.8%) - occurred between 35 and 39\nyears of age ( Table 2 ), but the groups were\nnot statistically different ( p =0.156).\nPregnancy rates and patient characteristics\nPregnancy rates according to maternal age\nEndometrial thickness >8mm was found in 76.6% (292) of the patients. Patients with\nan endometrial thickness greater than 8 mm were three times more likely to achieve\nclinical pregnancies than the individuals with an endometrial thickness <8mm\n( p =0.016). Of the 89 patients with endometrial thickness\n<8mm, only six (6.7%) achieved clinical pregnancies ( Table 1 ).\nFresh semen samples were more frequent (97.1%), and sperm concentration ranged from 2\nto 300 million per mL, while sperm motility ranged from 8.1% to 95% after\nprocessing. Cryopreserved semen was used in 11 procedures, and only one patient\nachieved clinical pregnancy. Motility above 55% had higher rates of clinical\npregnancy ( p =0.002).\nOf the 381 cycles analyzed, 328 (86.0%) were from Group 1 (injectables) ( Table 3 ), and 42 (11.0%) were from Group 2\n(injectables plus oral medication). Nine patients (2.3%) took only oral drugs and\ntwo (0.52%) did not use medication. The group given oral medication alone and the\nindividuals undergoing medication-free natural cycles did not achieve clinical\npregnancy.\nStimulation protocols and pregnancy\nIn the group taking only injectables, 159 patients (48.4%) took rFSH, 127 (38.7%)\nwere on hMG, and 42 (12.8%) on uFSH, yielding clinical pregnancy rates of 21.3%,\n10.4%, and 10.5%, respectively. Patients on rFSH achieved clinical pregnancy rates\n2.5 times higher than the individuals on hMG ( p =0.02) ( Table 4 ). Six (14.2%) of the 42 patients in\nGroup 2 achieved clinical pregnancies, three with clomiphene citrate and rFSH and\nthree with clomiphene citrate and hMG.\nPregnancy outcomes for each injectable drug used\n\nIt is important to point out that even in patients diagnosed with idiopathic\ninfertility, fertility rates may decline due to lower oocyte quality, fertilization\nfailure or embryo implantation abnormalities ( Wolff\n et al ., 2013 ). Advanced maternal age is an\nestablished independent negative prognostic factor for live births and clinical\npregnancy associated with lower chances of spontaneous or assisted pregnancy ( Geisler  et al ., 2017 ). In the\npresent study, a clear relationship was observed between lower clinical pregnancy\nrates and advanced maternal age. IUI procedures do not achieve high pregnancy rates\nin this group. In another similar study carried out in Brazil, advanced maternal age\nwas the only variable significantly correlated with success rates of IUI ( Sicchieri  et al ., 2018 ).\nAdequate endometrial thickness is widely considered a decisive factor in the\noutcomes of ART treatments ( Wolff et al.,\n2013 ). The finding in our study that more than 93% of the patients with\nendometrium thickness >8 mm did not achieve pregnancy corroborates other studies.\nIn a large study enrolling 2,929 patients submitted to IUI with idiopathic\ninfertility factor, the results plateaued at an endometrial thickness of 10 mm . The\nliterature is still controversial in relation to very thick endometria (>14mm),\nand some studies suggested that there might be less favorable endometrial\ncharacteristics in this group, as well as lower implantation rates ( Dietterich  et al ., 2002 ;  Weissman  et al ., 1999 ). Some\nstudies suggested that the ultrasound aspect of the endometrium (echogenicity) might\nbe linked to higher implantation rates. This was not analyzed in the present study\n( Alborzi  et al .,\n2005 ).\nThe link between increased pregnancy rates and increased sperm motility is in\nagreement with the literature ( Duran  et\nal ., 2003 ). Few studies have looked into the outcomes of IUI\nwith frozen semen. Our study corroborated the literature, suggesting lower pregnancy\nrates when frozen semen samples were used in IUI ( Dinelli  et al ., 2014 ). Since our study featured a\nlimited number of patients using frozen semen, more studies are needed to determine\nwhether IUI should be carried out with frozen semen. Since motility is very\nimportant to achieving successful outcomes, it has been suggested that the causes of\nlow sperm motility after semen processing, such as varicocele, should be corrected\nprior to assisted reproduction procedures ( Hendin\n et al ., 2000 ).\nAlthough only a few patients did not take injectables in our study, cycles stimulated\nwith injectable or oral medications and injectables only have reportedly greater\nchances of success. In a study by  Malchau  et\nal . (2014) , 76% of the children born after IUI resulted from\ncontrolled ovarian stimulation with CC, FSH or both ( Rashidi  et al ., 2013 ).\nA trend toward higher pregnancy rates was observed with rFSH ( Demirol & Gurgan, 2007 ), but the cost-effectiveness of this\ndrug compared to hMG is questionable, since rFSH is more expensive than hMG ( Gerli  et al ., 2008 ). In\naddition to providing better ovarian response, cycles with rFSH reportedly yield\nbetter quality oocytes ( Cheon  et\nal ., 2004 ). Our study showed better results in stimulated cycles\nusing rFSH, but the medication is not affordable by the standards of most of our\npatients. Moreover, positive pregnancy outcomes may also be achieved through other\nstimulation protocols.\n\nIUI is a viable procedure and a good option for the initial treatment of infertile\ncouples, since high pregnancy rates may be achieved when patients are adequately\nselected.\nIn this study, IUI patients younger than 40 years with endometrial thickness greater\nthan 8mm had higher clinical pregnancy rates. Endometrial thickness may be an\nimportant parameter to be assessed in future research. Sperm motility also had a\nsignificant impact on the success of IUI cycles. Further studies are needed to\nevaluate whether frozen sperm negatively affects pregnancy rates.\nOur results suggest that rFSH produces better outcomes than hMG, but more prospective\ncontrolled studies are needed to confirm the efficiency of these drugs in IUI.\nThe decision to perform the most appropriate treatment for each patient should be a\npriority in ART centers. It is important to carry out studies in each service and\ncompare the findings with data from the literature.","source_license":"CC-BY-4.0","license_restricted":false}