Reproduction, fertility, and development最新文献

{"title":"Betaine ameliorates high glucose-induced oxidative stress in granulosa cells.","authors":"Mohammad Hussein Abnosi,&nbsp;Mohammad Reza Tabandeh,&nbsp;Fatmeh Mosavi-Aroo","doi":"10.1071/RD22247","DOIUrl":"https://doi.org/10.1071/RD22247","url":null,"abstract":"<p><strong>Context: </strong>In diabetes, abnormalities of granulosa cells (GCs) and steroidogenesis are associated with hyperglycaemia-induced oxidative stress. Betaine has beneficial effect in experimental model of diabetes by reducing oxidative stress, inflammation, and apoptosis.</p><p><strong>Aims: </strong>In this study we investigate the effects of betaine to prevent oxidative stress in GCs induced by high glucose and improve steroidogenesis.</p><p><strong>Methods: </strong>Primary GCs, isolated from ovarian follicles of C57BL/6 mice were cultured in 5mM (control) and 30mM (hyperglycaemia) of glucose and in presence of 5mM of betaine for 24h. Then antioxidant enzymes, malondialdehyde, oestradiol and progesterone were measured. In addition, the expression of Nrf2 and NF-κB , antioxidant enzymes (Sod1 , Gpx and Cat ) were analysed by qRT-PCR assay.</p><p><strong>Key results: </strong>We observed significant (P <0.001) up-regulation of NF-κB and down-regulation of Nrf2 due to high concentration of glucose. Also significant (P <0.001) down-regulation of related antioxidant genes (Cat , Sod1 and GPx ) and activity reduction of these enzymes as well as significant (P <0.001) elevation of malondialdehyde was observed. In addition, betaine treatment compensated the drastic effect of high glucose induced oxidative stress via down-regulating the expression of NF-κB and up-regulating the expression of Nrf2 , Cat , Sod1 and GPx . It was also shown that betaine in the presence of FSH significantly (P <0.001) restored the oestradiol and progesterone level.</p><p><strong>Conclusion: </strong>Betaine compensated the antioxidant stress in mouse GCs under hyperglycaemic condition via regulation of Nrf2/NF-κB at transcription level.</p><p><strong>Implications: </strong>As betaine is a natural product and no side effect has been reported to today, we suggest more research needs to be carried out especially on patients whom suffer from diabetes to find the probability of using betaine as a therapeutic agent.</p>","PeriodicalId":20932,"journal":{"name":"Reproduction, fertility, and development","volume":"35 6","pages":"395-405"},"PeriodicalIF":1.9,"publicationDate":"2023-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9196223","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Quantitative proteomic analysis of heifer serum at the peri-implantation period using two complementary methods.","authors":"Jimena Inés Ruiz Álvarez,&nbsp;Juan Manuel Teijeiro,&nbsp;Caroline Evans,&nbsp;Patricia Estela Marini","doi":"10.1071/RD22228","DOIUrl":"https://doi.org/10.1071/RD22228","url":null,"abstract":"CONTEXT\u0000Systemic events resulting from mother-embryo relation at the peri-implantation period may result in specific changes to the protein composition of serum and thus provide a source of biomarkers for early detection of pregnancy.\u0000\u0000\u0000AIMS\u0000We set out to use two different quantitative proteomic approaches to test this hypothesis by comparing heifer serum at the peri-implantation period to that from cycling heifers.\u0000\u0000\u0000METHODS\u0000Two-dimensional electrophoresis (2-DE) and isobaric tags were used for relative and absolute quantitation (iTRAQ) of proteins.\u0000\u0000\u0000KEY RESULTS\u0000These methods yielded complementary data indicating biomarker candidate proteins.\u0000\u0000\u0000CONCLUSIONS\u0000Different proteomic methods provide different and complementary information that needs to be analysed in order to consider proteins as potential biomarkers.\u0000\u0000\u0000IMPLICATIONS\u0000In order to characterise the proteome under specific conditions, the use of complementary techniques is advisable.","PeriodicalId":20932,"journal":{"name":"Reproduction, fertility, and development","volume":"35 6","pages":"416-421"},"PeriodicalIF":1.9,"publicationDate":"2023-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9201415","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Knockdown of regulatory associated protein of TOR (raptor) in hypothalamus-stimulated folliculogenesis and induced ovarian cysts.","authors":"Pauline Tartarin,&nbsp;Matthieu Keller,&nbsp;Edith Guibert,&nbsp;Elliott Trives,&nbsp;Guillaume Bourdon,&nbsp;Pablo Chamero,&nbsp;Didier Negre,&nbsp;Fabien Cornilleau,&nbsp;Vanaique Guillory,&nbsp;Eric JeanPierre,&nbsp;Caroline Costa,&nbsp;Stéphanie Migrenne,&nbsp;Joelle Dupont,&nbsp;Pascal Froment","doi":"10.1071/RD21260","DOIUrl":"https://doi.org/10.1071/RD21260","url":null,"abstract":"<p><strong>Context: </strong>Mammalian target of rapamycin complex 1 (mTORC1) is an essential sensor that regulates fundamental biological processes like cell growth, proliferation and energy metabolism. The treatment of disease by sirolimus, a mTORC1 inhibitor, causes adverse effects, such as female fertility disorders.</p><p><strong>Aims: </strong>The objective of the study was to decipher the reproductive consequences of a downregulation of mTORC1 in the hypothalamus.</p><p><strong>Methods: </strong>The reduced expression of mTORC1 was induced after intracerebroventricular injection of lentivirus expressing a short hairpin RNA (shRNA) against regulatory associated protein of TOR (raptor) in adult female mice (ShRaptor mice).</p><p><strong>Key results: </strong>The ShRaptor mice were fertile and exhibited a 15% increase in the litter size compared with control mice. The histological analysis showed an increase in antral, preovulatory follicles and ovarian cysts. In the hypothalamus, the GnRH mRNA and FSH levels in ShRaptor mice were significantly elevated.</p><p><strong>Conclusions: </strong>These results support the hypothesis that mTORC1 in the central nervous system participates in the regulation of female fertility and ovarian function by influencing the GnRH neuronal activity.</p><p><strong>Implications: </strong>These results suggest that a lower mTORC1 activity directly the central nervous system leads to a deregulation in the oestrous cycle and an induction of ovarian cyst development.</p>","PeriodicalId":20932,"journal":{"name":"Reproduction, fertility, and development","volume":"35 4","pages":"307-320"},"PeriodicalIF":1.9,"publicationDate":"2023-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10765519","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Equine <i>in vitro</i> produced blastocysts: relationship of embryo morphology, stage and speed of development to foaling rate.","authors":"Niamh Lewis,&nbsp;Heloisa Canesin,&nbsp;Young Ho Choi,&nbsp;Rob Foss,&nbsp;Matheus Felix,&nbsp;Kindra Rader,&nbsp;Katrin Hinrichs","doi":"10.1071/RD22224","DOIUrl":"https://doi.org/10.1071/RD22224","url":null,"abstract":"<p><strong>Context: </strong>Information on factors associated with developmental competence of equine in vitro -produced (IVP) blastocysts is lacking.</p><p><strong>Aims: </strong>To determine the relationships of stage, grade, day of development, and specific morphological parameters of equine IVP blastocysts, to pregnancy and foaling rates.</p><p><strong>Methods: </strong>Photomicrographs of 316 IVP embryos with known pregnancy outcomes were scrutinised individually by four observers. Inter-observer variation was assessed, and pregnancy outcome evaluated in relation to day of blastocyst development and assigned grade and stage. Individual component analysis was performed to determine the association of specific morphological parameters with foaling rate.</p><p><strong>Key results: </strong>Overall pregnancy rate was 76.9% and foaling rate was 56.3%. The day of embryo development did not affect pregnancy rate but significantly affected foaling rate. Embryo stage did not affect foaling rate. Embryo grade affected foaling rate only for Day-9 embryos. Some morphological features in the bovine grading system did not predict outcome in equine IVP embryos. Significant individual parameters differed between Stage 5 and Stage 6 equine blastocysts.</p><p><strong>Conclusions: </strong>Day of blastocyst development is the major factor related to foaling rate for equine IVP embryos. Notably, there was no effect of embryo stage on foaling rate and no evidence that prolonging culture until embryos advance in stage increases foaling rate. The standard bovine grading system is not directly applicable to equine IVP embryos; equine-specific staging and grading systems are proposed.</p><p><strong>Implications: </strong>This information will allow laboratories to identify embryos with the highest developmental competence. Use of the proposed systems will increase consistency in embryo assessment among laboratories.</p>","PeriodicalId":20932,"journal":{"name":"Reproduction, fertility, and development","volume":"35 4","pages":"338-351"},"PeriodicalIF":1.9,"publicationDate":"2023-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10732201","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Ovarian and uterine changes during the oestrous cycle in female dogs.","authors":"Lena Lindh,&nbsp;Mariusz P Kowalewski,&nbsp;Anne-Rose Günzel-Apel,&nbsp;Sandra Goericke-Pesch,&nbsp;Vesa Myllys,&nbsp;Gerhard Schuler,&nbsp;Merja Dahlbom,&nbsp;Heli Lindeberg,&nbsp;Olli A T Peltoniemi","doi":"10.1071/RD22177","DOIUrl":"https://doi.org/10.1071/RD22177","url":null,"abstract":"<p><strong>Context: </strong>An accurate staging of sexual cycle is essential for the optimum timing of medical interventions.</p><p><strong>Aims: </strong>Here, an updated insight into clinical, endocrinological and vagino-cytological parameters, and their correlation with histomorphology of ovarian and uterine tissue samples is presented.</p><p><strong>Methods: </strong>Samples from 39 dogs were collected at various stages of the oestrous cycle: pro-oestrus (n =8), oestrus (n =12), dioestrus (n =9) (luteal phase) and anoestrus (n =10), according to clinical observations. Final allocation of samples was done after histomorphological evaluation of all tissues. Peripheral oestradiol-17ß (E2) and progesterone (P4) concentrations were measured, P4 by both chemiluminescent immunoassay (CLIA) and radioimmunoassay (RIA).</p><p><strong>Key results: </strong>Differences were observed between determination of the stage of the oestrous cycle, either by clinical, endocrinological or histomorphological evaluation. Individuals considered to be in clinical and endocrinological oestrus, had entered the luteal phase according to histomorphology. P4 concentrations measured by two different assays differed, underlying the importance to understand that absolute P4 concentrations may deviate depending on the used assay. Comparison of E2 and P4 concentrations is suggested to be useful when defining the transition from early follicular phase to the time of ovulation.</p><p><strong>Conclusions and implications: </strong>Based on parallel histomorphological observations, combined with clinical and endocrinological findings on the same individuals, the present study emphasises that an accurate classification of the stage of the cycle in female dogs based solely on clinical and endocrinological assessments can be difficult. The histomorphological findings presented herein provide new insights into the transitional phases between the different stages of the oestrous cycle in the dog.</p>","PeriodicalId":20932,"journal":{"name":"Reproduction, fertility, and development","volume":"35 4","pages":"321-337"},"PeriodicalIF":1.9,"publicationDate":"2023-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10786740","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Abstracts of the 1st International Noor-Al Hayat Fertility (INAF) Conference, 5–7 November 2022","authors":"","doi":"10.1071/rdv35n3abs","DOIUrl":"https://doi.org/10.1071/rdv35n3abs","url":null,"abstract":"","PeriodicalId":20932,"journal":{"name":"Reproduction, fertility, and development","volume":"1 1","pages":""},"PeriodicalIF":1.9,"publicationDate":"2023-01-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"46635882","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Sperm cryopreservation: current status and future developments.","authors":"Alena Hungerford,&nbsp;Hassan W Bakos,&nbsp;Robert John Aitken","doi":"10.1071/RD22219","DOIUrl":"https://doi.org/10.1071/RD22219","url":null,"abstract":"<p><p>The cryopreservation of spermatozoa is an important reproductive technology for the preservation of fertility in man and animals. Since the serendipitous discovery of glycerol as an effective cryoprotectant in 1947, sperm cryopreservation has undergone many changes in terms of the freezing methods employed, the rates at which samples are frozen and thawed, and the media used to preserve sperm functionality and DNA integrity. An extensive literature survey has been conducted addressing the cryoprotectants employed for both animal and human semen and the freezing protocols utilised. The results indicate that glycerol remains the dominant cryoprotective agent, usually incorporated into a balanced salt solution containing energy substrates, buffers, osmolytes and protein in the form of human serum albumin (human) or skimmed milk (animal). Realisation that some of the damage observed in cryostored cells involves the generation of reactive oxygen species during the thawing process, has prompted many studies to assess the relative merits of incorporating antioxidants into the cryopreservation media. However, in the absence of systematic comparisons, there is currently no consensus as to which antioxidant combination might be the most effective. Utilising our fundamental understanding of cryodamage to optimise cryopreservation protocols for each species will be important in the future.</p>","PeriodicalId":20932,"journal":{"name":"Reproduction, fertility, and development","volume":"35 3","pages":"265-281"},"PeriodicalIF":1.9,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10615345","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"\"Doing it on my own terms\": Transgender and nonbinary adults' experiences with HPV self-swabbing home testing kits.","authors":"Shanna K Kattari, Emma B Gross, Vern Harner, Emily Andrus, Daphna Stroumsa, Molly B Moravek, Andrew Brouwer","doi":"10.1080/23293691.2022.2094737","DOIUrl":"10.1080/23293691.2022.2094737","url":null,"abstract":"<p><p>The human papillomavirus (HPV) carries a significant health risk for people with a cervix. Among transgender and nonbinary people, however, testing and treatment for HPV can pose difficulties, and even be traumatic at times. This current study is part of a larger mixed methods study conducted in Michigan in 2020, and it explores the experiences of transmasculine and nonbinary people with at-home self-swabbing HPV test kits and knowledge of HPV transmission/screenings. Phenomenological methods were used by conducting virtual qualitative interviews with ten transmasculine and nonbinary individuals with cervixes, ages 23-59. Interviews were independently coded by members of the research team and a tabletop theming method was used. Four themes were generated from the data: 1) Multilevel barriers; 2) \"Get it done, so I know that I am safe\"; 3) Contrasting preferences for care; and 4) Community calls for change. The discussion focuses on the implications of these findings for improving sexual health care for the transgender and nonbinary community, along with directions for further research.</p>","PeriodicalId":20932,"journal":{"name":"Reproduction, fertility, and development","volume":"22 1","pages":"496-512"},"PeriodicalIF":0.0,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10720596/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"81815193","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Effect of melatonin and nitric oxide on capacitation and apoptotic changes induced by epidermal growth factor in ram sperm.","authors":"Sara Miguel-Jiménez,&nbsp;Melissa Carvajal-Serna,&nbsp;Victoria Peña-Delgado,&nbsp;Adriana Casao,&nbsp;Rosaura Pérez-Pe","doi":"10.1071/RD22146","DOIUrl":"https://doi.org/10.1071/RD22146","url":null,"abstract":"<p><strong>Context: </strong>Apart from the canonical cAMP-PKA pathway, ram sperm capacitation can be achieved by the MAPK ERK1/2 signalling cascade, activated by epidermal growth factor (EGF).</p><p><strong>Aims: </strong>This study aims to investigate the effect of melatonin and nitric oxide (NO·) on capacitation and apoptotic-like changes in EGF-capacitated ram spermatozoa.</p><p><strong>Methods: </strong>In vitro capacitation was induced by EGF in the absence or presence of melatonin (100pM or 1μM). Also, a NO· precursor, L-arginine, or a NOS inhibitor, NG-nitro-L-arginine methyl ester (L-NAME), were added to capacitation media to study the interaction of NO· and melatonin during EGF-capacitation. Sperm functionality parameters (motility, viability, capacitation state), apoptotic markers (caspase activation and DNA damage), NO· levels, and phosphorylated c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (assessed by Western blot), were evaluated in swim-up and capacitated samples with EGF.</p><p><strong>Key results: </strong>NO· levels and the apoptotic-related markers were raised after EGF incubation. Melatonin had a bimodal role on sperm EGF-capacitation, preventing it at high concentration and promoting acrosome reaction at low concentration, but neither of the two concentrations prevented the increase in apoptotic-like markers or NO· levels. However, melatonin at 1μM prevented the activation of JNK.</p><p><strong>Conclusions: </strong>NO· metabolism does not seem to modulate the apoptosis-like events in ram spermatozoa. Melatonin at 1μM prevents ram sperm capacitation induced by EGF independently from nitric oxide metabolism, and it could be exerted by limiting the JNK mitogen-activated protein kinase (MAPK) activation.</p><p><strong>Implications: </strong>This study improvesour understanding of the biochemical mechanisms involved in sperm capacitation, and ultimately, fertility.</p>","PeriodicalId":20932,"journal":{"name":"Reproduction, fertility, and development","volume":"35 3","pages":"282-293"},"PeriodicalIF":1.9,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10615300","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Persistent organic pollutants affect steroidogenic and apoptotic activities in granulosa cells and reactive oxygen species concentrations in oocytes in the mouse.","authors":"Kinga Krawczyk,&nbsp;Weronika Marynowicz,&nbsp;Karolina Pich,&nbsp;Oliwia Jędruch,&nbsp;Gabriela Kania,&nbsp;Justyna Gogola-Mruk,&nbsp;Wacław Tworzydło,&nbsp;Zbigniew Polański,&nbsp;Anna Ptak","doi":"10.1071/RD21326","DOIUrl":"https://doi.org/10.1071/RD21326","url":null,"abstract":"<p><strong>Context: </strong>The destruction of granulosa cells (GCs), the main functional cell type in the ovary, prevents steroid hormone production, which in turn may damage oocytes, resulting in ovarian failure. The accumulation of a number of persistent organic pollutants (POPs) in the ovarian follicular fluid (FF) has been documented, which raises serious questions regarding their impact on female fertility.</p><p><strong>Aims: </strong>We aimed to determine whether a mixture of POPs reflecting the profile found in FF influences mouse GCs or oocyte function and viability.</p><p><strong>Methods: </strong>A mixture of POPs, comprising perfluorooctanoate, perfluorooctane sulfonate, 2,2-dichlorodiphenyldichloroethylene, polychlorinated biphenyl 153, and hexachlorobenzene, was used. In addition to using the exact concentration of POPs previously measured in human FF, we tested two other mixtures, one with10-fold lower and another with 10-fold higher concentrations of each POP.</p><p><strong>Key results: </strong>Steroidogenesis was disrupted in GCs by the POP mixture, as demonstrated by lower oestradiol and progesterone secretion and greater lipid droplet accumulation. Furthermore, the POP mixture reduced GC viability and increased apoptosis, assessed using caspase-3 activity. The POP mixture significantly increased the number of oocytes that successfully progressed to the second meiotic metaphase and the oocyte reactive oxygen species (ROS) concentration.</p><p><strong>Conclusions: </strong>Thus, a mixture of POPs that are typically present in human FF has detrimental effects on ovarian function: it reduces the viability of GCs, and increases the oocyte concentrations of ROS.</p><p><strong>Implications: </strong>These results indicate that chronic exposure to POPs adversely affects female reproductive health.</p>","PeriodicalId":20932,"journal":{"name":"Reproduction, fertility, and development","volume":"35 3","pages":"294-305"},"PeriodicalIF":1.9,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10615299","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}