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1.
J Assist Reprod Genet ; 39(4): 945-952, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-35338418

RESUMO

PURPOSE: To test the validity of the Vienna consensus laboratory key performance indicators (KPIs) to monitor the outcome of treatments involving women of different age ranges. METHODS: The retrospective cohort study included 862 complete IVF/ICSI cycles carried out between January 2014 and May 2021. All embryos of each cycle cohort were subject to extended culture. The overall population was divided into two groups according to female age: the Vienna consensus (≤ 39 years) and older female age (≥ 40 years). We compared outcomes of a selection of the Vienna performance indicators (PIs) and KPIs, with a focus on measures relevant to embryo cleavage and blastocyst formation. A possible association between total good blastocyst development rate (TGBDR) and cumulative clinical pregnancy rate (CPR) was also assessed. RESULTS: No differences were observed in fertilization and embryo cleavage KPIs between the Vienna consensus and the older female age group (standard IVF fertilization, 67.2 vs. 67.3; ICSI fertilization, 72.3 vs. 75.3; day 2 development, 57.6% vs 58.7%; day 3 development, 52.4% vs. 50.7%, respectively). TGBDR was lower in the older female age group (45.5% vs. 33.4% p < 0.001). Multivariate logistic regression analysis indicated female age as a factor independently associated with TGBDR. Clinical outcomes significantly decreased with increasing female age. CONCLUSION: The study suggests that, while most laboratory outcome measures are reliably applicable irrespective of female age, KPIs describing extended embryo culture should be fine-tuned in consideration of older female age.


Assuntos
Blastocisto , Fertilização in vitro , Adulto , Consenso , Feminino , Humanos , Gravidez , Taxa de Gravidez , Estudos Retrospectivos
2.
J Reprod Infertil ; 22(3): 159-164, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34900636

RESUMO

BACKGROUND: Despite a plethora of studies conducted so far, a debate is still unresolved as to whether TLM can identify predictive kinetic biomarkers or algorithms universally applicable. Therefore, this study aimed to elucidate if there is a relationship between kinetic variables and ploidy status of human embryos or blastocyst developmental potential. METHODS: For conducting this retrospective cohort study, the normal distribution of data was verified using Kolmogorov-Smirnov test with the Lilliefors' amendment and the Shapiro-Wilk test. Kinetic variables were expressed as median and quartiles (Q1, Q2, Q3, Q4). Mann-Whitney U-test was used to compare the median values of parameters. Univariate and multiple logistic regression models were used to assess relationship between blastocyst developmental potential or ploidy status and kinetics. Several confounding factors were also assessed. RESULTS: Blastocyst developmental potential was positively correlated with the t4-t3 interval (s2) (OR=1.417, 95% CI of 1.288-1.560). s2 median value was significantly different between high- and low-quality blastocysts (0.50 and 1.33 hours post-insemination, hpi, respectively; p=0.003). In addition, timing of pronuclear appearance (tPNa) (OR=1.287; 95% CI of 1.131-1.463) had a significant relationship with ploidy changes. The median value of tPNa was statistically different (p=0.03) between euploid and aneuploid blastocysts (Euploid blastocysts=8.9 hpi; aneuploid blastocysts=10.3 hpi). CONCLUSION: The present findings are in line with the study hypothesis that kinetic analysis may reveal associations between cleavage patterns and embryo development to the blastocyst stage and ploidy status.

3.
Mol Hum Reprod ; 27(7)2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-34191013

RESUMO

The sperm is essential for reconstitution of embryonic diploidy and highly specialized developmental functions. Immediately after gamete fusion, the sperm-borne PLC-zeta triggers activation, generating intracellular free Ca2+ oscillations. Mutations in the PLC-zeta encoding gene are associated with the absence of this factor in mature sperm and inability to achieve fertilization. Sperm play also a role in the greater game of the choreography of fertilization. In the human, the sperm centrioles are introduced into the oocyte environment with gamete fusion. They interact with the oocyte cytoskeletal apparatus to form a functional pair of centrosomes and ultimately regulate pronuclear juxtaposition in preparation for the first cleavage. As a consequence, the fidelity of chromosome segregation during the first cell divisions depends on the function of sperm centrioles. Sperm DNA integrity is essential for embryo development and health. Damaged DNA does not impact on the sperm fertilization ability following ICSI. However, detrimental effects emerge at pre- and post-implantation stages. Sperm-specific epigenetic factors also play an active role in the regulation of embryonic development, as shown by correlations between reduced embryo morphological quality and incorrect chromatin packaging during spermiogenesis or abnormal methylation of sperm CpG islands. This functional landscape demonstrates that the contribution of the sperm to development goes far beyond its well-established role in fertilization. Clinical studies confirm this view and indicate sperm function as a crucial aspect of research to increase the efficacy of assisted reproduction treatments.


Assuntos
Desenvolvimento Embrionário , Espermatozoides/fisiologia , Aneuploidia , Animais , Blastocisto/metabolismo , Sinalização do Cálcio , Centríolos/fisiologia , Cromatina/ultraestrutura , Ilhas de CpG , Fragmentação do DNA , Metilação de DNA , Desenvolvimento Embrionário/genética , Feminino , Fertilização , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Masculino , Fosfoinositídeo Fosfolipase C/fisiologia , Gravidez , Resultado da Gravidez , RNA/genética , Técnicas de Reprodução Assistida , Interações Espermatozoide-Óvulo , Espermatozoides/enzimologia
4.
J Assist Reprod Genet ; 38(7): 1737-1743, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33821429

RESUMO

PURPOSE: To study embryo morphokinetics in relation to release in spent media of molecules with possible roles in development and implantation (miR-20a, miR-30c, and sHLA-G). METHODS: Data were obtained from embryos generated in standard IVF and ICSI cycles. The Eeva system was used for embryo assessment, based on early morphokinetic parameters and producing a score (1-5, best-worst) corresponding to higher/medium/lower chances of development to blastocyst. miRNAs - mm miR-20a-5p and miR-30c-5p - and sHLA-G were quantified in 25 µl of spent blastocyst media (SBM) collected before vitrification or transfer. Statistical analyses were performed applying Kolmogorov-Smirnov, Shapiro-Wilk, and Spearman's correlation coefficient tests, where appropriate. RESULTS: SBM were collected from a total of 172 viable blastocysts. Their analysis showed that concentration of miR-20a was progressively lower as Eeva score increased and probability of development to blastocyst decreased (P = 0.016). The opposite trend was observed in the case of miR-30c, i.e., concentration was higher as score increased and chances of development to blastocyst decreased (P = 0.004). Analysis of sHLA-G revealed a negative correlation with Eeva score, i.e., levels were progressively lower as Eeva score increased and probability of development to blastocyst decreased (R = - 0.388, N = 141, P = 0.001). CONCLUSION: Our data suggest that morphokinetic algorithms that predict development to blastocyst stage, in fact, also identify embryos with molecular and cellular profiles more consistent with developmental functions.


Assuntos
Blastocisto/fisiologia , Implantação do Embrião/fisiologia , Antígenos HLA-G/análise , MicroRNAs/análise , Adulto , Biomarcadores/análise , Blastocisto/citologia , Substitutos Ósseos , Meios de Cultura/análise , Técnicas de Cultura Embrionária/métodos , Feminino , Fertilização in vitro , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Masculino , Estudo de Prova de Conceito
5.
J Assist Reprod Genet ; 37(7): 1653-1659, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32405898

RESUMO

PURPOSE: To explore the possible influence of sperm quality, as assessed by prewash total sperm count (TSC), on cumulative success rates in assisted reproduction cycles. METHODS: Retrospective study carried out in private IVF centre. Seven hundred sixty-five couples undergoing complete ICSI cycles, i.e. whose all embryos were transferred or disposed of. Couples were characterised by male infertility and female age younger than 36 years. Couples with a combination of female and male infertility factors were excluded. The primary outcome measure was cumulative live birth rate. Secondary outcomes were cumulative pregnancy and miscarriage rates. No specific interventions were made. RESULTS: Higher TSC values have a positive impact on cumulative success rates in cycles characterised by few retrieved oocytes (1 to 5), while does not influence the outcome of cycles with a normal (6 to 10) or high (> 10) number of retrieved oocytes. CONCLUSIONS: The study highlights the importance of sperm quality for the efficacy of assisted reproduction treatments. This influence may remain relatively cryptic in association with normal or high ovarian response, but emerge decisively in cases of reduced ovarian response, suggesting a relationship between ovarian response and oocyte ability to compensate for paternal-derived deficiencies.


Assuntos
Infertilidade Masculina , Contagem de Espermatozoides , Injeções de Esperma Intracitoplásmicas/métodos , Adulto , Coeficiente de Natalidade , Feminino , Humanos , Infertilidade Masculina/terapia , Nascido Vivo , Masculino , Recuperação de Oócitos , Indução da Ovulação , Gravidez , Taxa de Gravidez , Estudos Retrospectivos , Injeções de Esperma Intracitoplásmicas/estatística & dados numéricos , Interações Espermatozoide-Óvulo/fisiologia , Resultado do Tratamento
6.
J Assist Reprod Genet ; 36(10): 2047-2055, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31630313

RESUMO

PURPOSE: In this study, we tested the hypothesis that, in PGT-A cycles, decreased semen quality is associated with increased rates of mosaic blastocysts. METHODS: In a retrospective analysis, three hundred and forty PGT-A cycles are divided into study groups according to semen quality. Cycles were initially divided into two groups, discerning couples with absence of male factor of infertility (non-male factor: NMF; N = 146 cycles) from couples with a male factor of infertility (MF; N = 173 cycles). Couples with severe male factor (SMF) infertility (n = 22) were assessed separately. Embryos were cultured to the blastocyst stage and chromosomally assessed by array comparative genomic hybridization (aCGH). The study did not involve specific interventions. RESULTS: The reproductive outcome of MF and NMF groups did not indicate statistically significant differences. However, while no differences were found between MF and NMF groups in terms of euploid or aneuploid blastocysts rates, a significantly higher rate of mosaic blastocysts was observed in the MF group (3.6% vs. 0.5%, respectively; P = 0.03). A similar pattern of results was observed in the SMF group when compared with those of the other PGT-A cycles taken together (no SMF). In particular, a significantly higher rate of mosaic blastocysts was observed in the SMF group (7.7% and 1.8%, respectively; P = 0.008). CONCLUSIONS: The study outcome strongly suggests that compromised semen quality is associated with increased rates of mosaic blastocysts analysed in PGT-A cycles. Sperm assessment appears therefore as an important factor in the determination of embryo development and for a more precise prognostic assessment of PGT-A cases.


Assuntos
Blastocisto/fisiologia , Infertilidade Masculina/genética , Infertilidade Masculina/fisiopatologia , Adulto , Aneuploidia , Hibridização Genômica Comparativa/métodos , Técnicas de Cultura Embrionária/métodos , Implantação do Embrião/genética , Transferência Embrionária/métodos , Desenvolvimento Embrionário/genética , Feminino , Fertilização in vitro/métodos , Testes Genéticos/métodos , Humanos , Masculino , Pessoa de Meia-Idade , Gravidez , Taxa de Gravidez , Diagnóstico Pré-Implantação/métodos , Estudos Retrospectivos , Análise do Sêmen/métodos
7.
Adv Exp Med Biol ; 1166: 169-187, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31301052

RESUMO

In the female reproductive tract, male gametes undergo a natural sperm selection process in order to discriminate spermatozoa on the basis of their quality to maximize the chances of successful reproduction. With the introduction of assisted reproductive technology (ART), scientists and clinicians developed diverse sperm selection strategies focusing on the isolation of competent spermatozoa. With increasing understanding of sperm functions and fertilization mechanism and evolution of available technologies, the initial simple sperm preparation protocols were complemented, and sometimes replaced, by new sperm-sorting techniques. In particular, while in the early years the focus was on obtaining motile spermatozoa, in later years, especially after the introduction of intracytoplasmic sperm injection (ICSI), the focus shifted to the isolation of functional and "healthy" spermatozoa, considering some other important factors, such as sperm DNA integrity. Sperm DNA damage, as well as chromatin structure alterations, in fact, is related to decreased reproductive ability of men, in natural as well as in assisted reproduction.


Assuntos
DNA/normas , Técnicas de Reprodução Assistida , Espermatozoides , Dano ao DNA , Humanos , Masculino , Injeções de Esperma Intracitoplásmicas
9.
Medicine (Baltimore) ; 95(20): e3624, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-27196465

RESUMO

Predicting the outcome of in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) is one main goal of the present research on assisted reproduction. To understand whether density gradient centrifugation (DGC), used to select sperm, can affect sperm DNA integrity and impact pregnancy rate (PR), we prospectively evaluated sperm DNA fragmentation (sDF) by TUNEL/PI, before and after DGC. sDF was studied in a cohort of 90 infertile couples the same day of IVF/ICSI treatment. After DGC, sDF increased in 41 samples (Group A, median sDF value: 29.25% [interquartile range, IQR: 16.01-41.63] in pre- and 60.40% [IQR: 32.92-93.53] in post-DGC) and decreased in 49 (Group B, median sDF value: 18.84% [IQR: 13.70-35.47] in pre- and 8.98% [IQR: 6.24-15.58] in post-DGC). PR was 17.1% and 34.4% in Group A and B, respectively (odds ratio [OR]: 2.58, 95% confidence interval [CI]: 0.95-7.04, P = 0.056). After adjustment for female factor, female and male age and female BMI, the estimated OR increased to 3.12 (95% CI: 1.05-9.27, P = 0.041). According to the subgroup analysis for presence/absence of female factor, heterogeneity in the association between the Group A and B and PR emerged (OR: 4.22, 95% CI: 1.16-15.30 and OR: 1.53, 95% CI: 0.23-10.40, respectively, for couples without, n = 59, and with, n = 31, female factor).This study provides the first evidence that the DGC procedure produces an increase in sDF in about half of the subjects undergoing IVF/ICSI, who then show a much lower probability of pregnancy, raising concerns about the safety of this selection procedure. Evaluation of sDF before and after DGC configures as a possible new prognostic parameter of pregnancy outcome in IVF/ICSI. Alternative sperm selection strategies are recommended for those subjects who undergo the damage after DGC.


Assuntos
Centrifugação com Gradiente de Concentração , Fragmentação do DNA , Taxa de Gravidez , Espermatozoides , Adulto , Feminino , Humanos , Infertilidade Feminina/terapia , Infertilidade Masculina/terapia , Masculino , Gravidez , Estudos Prospectivos , Curva ROC , Injeções de Esperma Intracitoplásmicas
10.
J Assist Reprod Genet ; 31(8): 1045-51, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24906302

RESUMO

PURPOSE: To investigate whether the sperm fertilizing potential can be improved by selecting a non-apoptotic fraction using magnetic activated cell sorting (MACS), and to compare the results with the conventional swim-up method. METHODS: Twenty five male patients attending the andrology laboratory for sperm DNA fragmentation analysis. The sperm were prepared by density gradient centrifugation (DGC) and subsequently divided into three aliquots. The first was further separated into Annexin V-negative (non-apoptotic) fraction using MACS, the second was further processed by swim-up, while the third was left unseparated as a control. The impact of the combination of DGC with the two sperm preparation techniques on sperm quality was evaluated by comparing 'rapid progressive' motility, normal morphology according to Tygerberg's strict criteria and DNA integrity (by terminal deoxynucleotide transferase-mediated dUTP nick-end labeling [TUNEL]) for each aliquot. RESULTS: Sperm preparation that combines DGC with conventional swim-up method can provide sperm of higher quality in terms of motility, morphology and extent of DNA fragmentation compared to the Annexin V-negative (non-apoptotic) fraction derived from the combination of DGC with MACS. CONCLUSIONS: Integrating MACS as a part of sperm preparation technique will not improve sperm fertilizing potential to the same extent as the traditional swim-up separation procedure.


Assuntos
Anexina A5/metabolismo , Apoptose , Separação Celular/métodos , Magnetismo , Técnicas de Reprodução Assistida , Motilidade dos Espermatozoides , Espermatozoides/citologia , Adulto , Citometria de Fluxo , Humanos , Infertilidade Masculina , Masculino , Análise do Sêmen
11.
Adv Urol ; 2012: 854837, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22194740

RESUMO

Cryopreservation of human spermatozoa-introduced in the 1960's-has been recognized as an efficient procedure for management of male fertility before therapy for malignant diseases, vasectomy or surgical infertility treatments, to store donor and partner spermatozoa before assisted reproduction treatments and to ensure the recovery of a small number of spermatozoa in severe male factor infertility. Despite the usefulness of it, cryopreservation may lead to deleterious changes of sperm structure and function: while the effects of cryopreservation on cells are well documented, to date there is no agreement in the literature on whether or not cryopreservation affects sperm chromatin integrity or on the use of a unique and functional protocol for the freezing-thawing procedure. Therefore, sperm cryopreservation is an important component of fertility management and much of its successful application seems to affect the reproductive outcome of assisted reproduction technologies (ART): appropriate use of cryoprotectants before and sperm selection technologies after cryopreservation seem to have the greatest impact on preventing DNA fragmentation, thus improving sperm cryosurvival rates.

12.
Reprod Biomed Online ; 19 Suppl 3: 35-43, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-20034422

RESUMO

Hyaluronan has recently been employed in the development of a commercial diagnostic kit for assessing sperm maturity, the so-called sperm-hyaluronan-binding assay (HBA). The aim of this study was to evaluate the usefulness of this test, in addition to routine semen analysis, to identify patients with poor reproductive prognosis in conventional IVF. Furthermore, the ability of hyaluronan to select spermatozoa with low DNA fragmentation was investigated. A total of 60 IVF patients were analysed with regard to reproductive outcome, sperm parameters, HBA score and sperm DNA fragmentation. The DNA fragmentation analysis was performed using the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling assay on the total sperm population and on the hyaluronan-bound spermatozoa obtained from the same samples. No relationship between hyaluronan binding and fertilization, cleavage, good-quality embryos, implantation, clinical pregnancy, miscarriages and biochemical pregnancy rates was found. Otherwise, correlations between spermatozoa hyaluronan binding and morphology (P < 0.01) and a significant difference between DNA fragmentation of the total sperm population and DNA fragmentation of the hyaluronan-bound spermatozoa (P = 0.029) were found. The results underline the ability of hyaluronan to select spermatozoa with higher DNA integrity and morphology. Nevertheless, the clinical value of the HBA in the management of male infertility seems to be limited.


Assuntos
Fertilização in vitro/métodos , Ácido Hialurônico , Maturação do Esperma/fisiologia , Adulto , Distribuição de Qui-Quadrado , Fragmentação do DNA , Transferência Embrionária , Feminino , Humanos , Marcação In Situ das Extremidades Cortadas , Nascido Vivo , Masculino , Recuperação de Oócitos , Indução da Ovulação , Gravidez , Taxa de Gravidez , Contagem de Espermatozoides , Motilidade dos Espermatozoides
13.
Reprod Biomed Online ; 19 Suppl 3: 45-55, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-20034423

RESUMO

To date, several publications have focused their attention on a new method for observing spermatozoa called 'motile sperm organelle morphology examination' (MSOME), which enables the evaluation of the fine nuclear morphology of motile spermatozoa in real time at high magnification (>x6000). As a consequence, a new microinjection procedure called intracytoplasmic morphologically selected sperm injection (IMSI) has been developed. The aim of the present work is therefore to evaluate the efficacy of the IMSI technique in the light of the current literature, focusing attention on the potential clinical application of the selection of strictly morphologically normal spermatozoa in patients undergoing conventional intracytoplasmic sperm injection treatments. In addition, a brief analysis of preliminary data regarding the relationship between IMSI and assisted reproduction treatment outcome is presented.


Assuntos
Forma Celular/fisiologia , Injeções de Esperma Intracitoplásmicas/métodos , Espermatozoides/citologia , Feminino , Humanos , Infertilidade , Masculino , Microinjeções , Gravidez , Taxa de Gravidez , Motilidade dos Espermatozoides/fisiologia , Resultado do Tratamento
14.
Reprod Biomed Online ; 18(4): 486-95, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19400989

RESUMO

Sperm protamine deficiency and DNA damage were analysed employing chromomycin A(3) (CMA(3)) staining and the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling assay, respectively, in 132 patients (82 IVF, 50 intracytoplasmic sperm injection [ICSI]). The antioxidant ability of seminal plasma was analysed in 10 men, using the total oxidant scavenging capacity assay. A significant negative correlation was found between abnormal protamination and sperm parameters, including sperm DNA fragmentation (P < 0.01). A close relationship was found between sperm protamination and fertilization and pregnancy only in IVF (P = 0.004 and P < 0.04, respectively); in ICSI there was a correlation between DNA fragmentation and pregnancy (P = 0.031). Finally, there was a negative correlation between chromatin under-protamination and the antioxidant ability of seminal plasma (P < 0.01). Results of this study underline that, despite sperm abnormal protamination and DNA fragmentation being positively correlated, they affect the reproductive outcome in different ways: in particular there was good prognostic value for CMA(3) analysis only in IVF, whereas DNA fragmentation analysis was prognostic only for ICSI outcome. Data are also provided to support the idea of a relationship between defective antioxidant system activity and impairment of chromatin packaging.


Assuntos
Montagem e Desmontagem da Cromatina/fisiologia , Fragmentação do DNA , Fertilização in vitro/métodos , Protaminas/análise , Análise do Sêmen/métodos , Injeções de Esperma Intracitoplásmicas/métodos , Espermatozoides/citologia , Adulto , Cromomicina A3 , Feminino , Humanos , Marcação In Situ das Extremidades Cortadas , Itália , Masculino , Gravidez , Resultado da Gravidez , Espermatozoides/química
15.
Hum Reprod ; 20(8): 2140-52, 2005 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-15845594

RESUMO

BACKGROUND: A protocol for the chromosomal analysis of sperm samples with a severely reduced number of sperm cells was designed. METHODS: A severe male factor condition was the main cause of infertility for 38 couples: 27 were oligoasthenoteratospermic (OAT) and 11 with non-obstructive azoospermia underwent testicular sperm extraction (TESE). A two-round fluorescence in situ hybridization (FISH) protocol was performed with probes specific for the chromosomes X, Y, 13, 15, 16, 17, 18, 21 and 22. The recording of the position of each sperm cell at the microscope allowed diagnosis of each spermatozoon for the nine tested chromosomes. RESULTS: A mean number of 122+/-78.5 sperm were diagnosed per patient with an incidence of total abnormalities corresponding to 13.4%. chi2-tests for the observed frequencies and goodness-of-fit test were highly significant in all cases. A significantly higher proportion of total aneuploidy was detected in 79% of the tested samples compared to the normal population. Testicular sperm were significantly more prone to aneuploidy than ejaculated sperm. CONCLUSIONS: The designed FISH protocol for the analysis of severe OAT and TESE sperm samples is reliable, implying that the studied sample is representative of the original population. In view of the high incidence of aneuploidy in most severe OAT and TESE sperm, the FISH analysis of pathological sperm samples can be routinely performed in order to estimate the chances of the paternal contribution to aneuploidy in the resulting embryos.


Assuntos
Aneuploidia , Oligospermia/diagnóstico , Oligospermia/genética , Diagnóstico Pré-Implantação/métodos , Espermatozoides , Adulto , Feminino , Humanos , Hibridização in Situ Fluorescente , Incidência , Masculino , Pessoa de Meia-Idade , Oligospermia/epidemiologia , Diagnóstico Pré-Implantação/normas , Reprodutibilidade dos Testes , Técnicas de Reprodução Assistida
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