{"id":{"repo_id":"stellenbosch","oai_identifier":"oai:scholar.sun.ac.za:10019.1/135707"},"canonical_url":"https://search.dev.ndltd.org/etd/stellenbosch/oai:scholar.sun.ac.za:10019.1/135707","repository":{"repo_id":"stellenbosch","name":"Stellenbosch University","base_url":"https://scholar.sun.ac.za/server/oai/request"},"display":{"title":"Obesity-Induced Alterations in Sperm Quality, Testicular Morphology, and Extracellular Vesicle Localisation in Male Mice","abstract":"Background: Infertility is a recognised reproductive health issue affecting individuals worldwide. An increase in male infertility has been reported, with concurrent evidence of declining semen quality. Obesity is a major contributor to infertility, with obese men presenting with decreased testosterone levels, impaired spermatogenesis and reduced sperm function. Extracellular vesicles (EVs), which facilitate cell-to-cell communication, have been discovered in various biological fluids. Specifically, in the male reproductive system, testicular-derived EVs have been reported to regulate testosterone production and spermatogenesis. There is a significant gap in knowledge regarding the effects of obesity on testicular EVs and their potential role in altering the testicular microenvironment. Therefore, examining testicular-derived EVs may provide an understanding of the association between obesity and male infertility. Methods: This study utilised a genetic mouse model of obesity (ob/ob) to investigate obesity-induced male infertility. Control (C57BL/6) and obese (ob/ob) male mice were euthanized at 16 weeks of age. Epididymides were harvested to extract sperm to measure sperm concentration, motility (total, progressive and non-progressive), kinematics (straight line velocity (VSL), curvilinear velocity (VCL), amplitude of lateral head displacement (ALH) and linearity (LIN)), and morphology via Computer-Aided Sperm Analysis (CASA). Additionally, the testes were harvested and fixed in formalin or frozen for subsequent analyses. Formalin-fixed testes underwent haematoxylin and eosin (H&E), immunohistochemistry (IHC) and immunofluorescent (IF) staining to investigate testicular morphology and assessment of EV localisation, respectively. EV-specific biomarkers such as CD63, CD9 and ANXA1 were targeted. Frozen testes were utilised for EV extraction and further western blotting (WB) analyses. Spermatogenesis was investigated via WB for spermatogenesis-related proteins (IGF1, INSL6, and PRDX2). For EV extraction, frozen testes were dissociated and underwent differential centrifugation after which WB was performed to confirm EV isolation by targeting EV-specific biomarkers. Results: At 16-weeks old, obese mice had significantly increased body weights (P<0.0001) and fasting blood glucose levels (P=0.0004). Sperm concentration remained unchanged between groups. Obese spermatozoa had significantly reduced total (P<0.0001), progressive (P=0.009) and non-progressive (P<0.0001) motility when compared to controls. Kinematic measurements indicated reduced VSL (P=0.0064), VCL (P<0.0001) and ALH (P<0.0001) parameters for obese sperm, with LIN remaining unchanged. Obese mice presented with significantly increased abnormal spermatozoa (P<0.0001) and showed significant increased head and midpiece defects (P<0.0001 and P=0.0089, respectively). Obese testicular morphology displayed decreased seminiferous tubular area (P=0.0287) and diameter (P=0.0252), decreased lumen diameter (P=0.0011), as well as a high presence of vacuolation and disorganised cellular arrangements. IHC staining showed visible differences in EV localisation between control and obese testes for CD63, CD9 and ANXA1, with IF staining showing a decreased mean fluorescent intensity for CD63 (P=0.0345) in obese testes compared to controls. WB for IGF1, INSL6 and PRDX2 remained unchanged. WB for extracted EVs (CD63, CD9 and ANXA1) could not be quantified. Conclusion: Results from this study showed obesity-induced alterations in sperm parameters. Additionally, the testes and EV localisation results suggest a significant difference in CD63 EV marker compared to controls, suggesting less EVs in the ob/ob mice. Further investigation is required to delve deeper into the association between obesity, EV dynamics and testicular functions.","abstract_html":"Background: Infertility is a recognised reproductive health issue affecting individuals worldwide. An increase in male infertility has been reported, with concurrent evidence of declining semen quality. Obesity is a major contributor to infertility, with obese men presenting with decreased testosterone levels, impaired spermatogenesis and reduced sperm function. Extracellular vesicles (EVs), which facilitate cell-to-cell communication, have been discovered in various biological fluids. Specifically, in the male reproductive system, testicular-derived EVs have been reported to regulate testosterone production and spermatogenesis. There is a significant gap in knowledge regarding the effects of obesity on testicular EVs and their potential role in altering the testicular microenvironment. Therefore, examining testicular-derived EVs may provide an understanding of the association between obesity and male infertility. Methods: This study utilised a genetic mouse model of obesity (ob/ob) to investigate obesity-induced male infertility. Control (C57BL/6) and obese (ob/ob) male mice were euthanized at 16 weeks of age. Epididymides were harvested to extract sperm to measure sperm concentration, motility (total, progressive and non-progressive), kinematics (straight line velocity (VSL), curvilinear velocity (VCL), amplitude of lateral head displacement (ALH) and linearity (LIN)), and morphology via Computer-Aided Sperm Analysis (CASA). Additionally, the testes were harvested and fixed in formalin or frozen for subsequent analyses. Formalin-fixed testes underwent haematoxylin and eosin (H&amp;E), immunohistochemistry (IHC) and immunofluorescent (IF) staining to investigate testicular morphology and assessment of EV localisation, respectively. EV-specific biomarkers such as CD63, CD9 and ANXA1 were targeted. Frozen testes were utilised for EV extraction and further western blotting (WB) analyses. Spermatogenesis was investigated via WB for spermatogenesis-related proteins (IGF1, INSL6, and PRDX2). For EV extraction, frozen testes were dissociated and underwent differential centrifugation after which WB was performed to confirm EV isolation by targeting EV-specific biomarkers. Results: At 16-weeks old, obese mice had significantly increased body weights (P&lt;0.0001) and fasting blood glucose levels (P=0.0004). Sperm concentration remained unchanged between groups. Obese spermatozoa had significantly reduced total (P&lt;0.0001), progressive (P=0.009) and non-progressive (P&lt;0.0001) motility when compared to controls. Kinematic measurements indicated reduced VSL (P=0.0064), VCL (P&lt;0.0001) and ALH (P&lt;0.0001) parameters for obese sperm, with LIN remaining unchanged. Obese mice presented with significantly increased abnormal spermatozoa (P&lt;0.0001) and showed significant increased head and midpiece defects (P&lt;0.0001 and P=0.0089, respectively). Obese testicular morphology displayed decreased seminiferous tubular area (P=0.0287) and diameter (P=0.0252), decreased lumen diameter (P=0.0011), as well as a high presence of vacuolation and disorganised cellular arrangements. IHC staining showed visible differences in EV localisation between control and obese testes for CD63, CD9 and ANXA1, with IF staining showing a decreased mean fluorescent intensity for CD63 (P=0.0345) in obese testes compared to controls. WB for IGF1, INSL6 and PRDX2 remained unchanged. WB for extracted EVs (CD63, CD9 and ANXA1) could not be quantified. Conclusion: Results from this study showed obesity-induced alterations in sperm parameters. Additionally, the testes and EV localisation results suggest a significant difference in CD63 EV marker compared to controls, suggesting less EVs in the ob/ob mice. Further investigation is required to delve deeper into the association between obesity, EV dynamics and testicular functions.","abstract_has_math":false,"creators":["Uys, Sharnah"],"institution":"Stellenbosch : Stellenbosch University","degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":["Skosana, Bongekile Trisha","Marais, Erna"],"committee_chairs":[],"committee_members":[],"year":2026,"date_issued":"2026-03","date_published":"2026-03","updated_at":"2026-07-24T04:40:12Z","subjects":[],"languages":["en"],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"https://scholar.sun.ac.za/handle/10019.1/135707","outbound_label":"Repository record","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.advisor","label":"Advisor","values":["Skosana, Bongekile Trisha","Marais, Erna"]},{"key":"dc:contributor.other","label":"Dc Contributor Other","values":["Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Division of Medical Physiology."]},{"key":"dc:creator","label":"Author","values":["Uys, Sharnah"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.accessioned","label":"Dc Date Accessioned","values":["2026-04-08T09:54:59Z"]},{"key":"dc:date.available","label":"Dc Date Available","values":["2026-04-08T09:54:59Z"]},{"key":"dc:date.issued","label":"Date","values":["2026-03"]},{"key":"dc:publisher","label":"Institution","values":["Stellenbosch : Stellenbosch University"]},{"key":"dc:type","label":"Dc Type","values":["Thesis"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language.iso","label":"Language (ISO)","values":["en"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.uri","label":"Identifier URI","values":["https://scholar.sun.ac.za/handle/10019.1/135707"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["Thesis (MSc)--Stellenbosch University, 2026.","Uys, S. 2026. Obesity-Induced Alterations in Sperm Quality, Testicular Morphology, and Extracellular Vesicle Localisation in Male Mice. Unpublished masters thesis. Stellenbosch: Stellenbosch University [online]. Available: https://scholar.sun.ac.za/items/cf57fd1c-cc36-4e58-b690-6f5273ad443a"]},{"key":"dc:description.abstract","label":"Abstract","values":["Background: Infertility is a recognised reproductive health issue affecting individuals worldwide. An increase in male infertility has been reported, with concurrent evidence of declining semen quality. Obesity is a major contributor to infertility, with obese men presenting with decreased testosterone levels, impaired spermatogenesis and reduced sperm function. Extracellular vesicles (EVs), which facilitate cell-to-cell communication, have been discovered in various biological fluids. Specifically, in the male reproductive system, testicular-derived EVs have been reported to regulate testosterone production and spermatogenesis. There is a significant gap in knowledge regarding the effects of obesity on testicular EVs and their potential role in altering the testicular microenvironment. Therefore, examining testicular-derived EVs may provide an understanding of the association between obesity and male infertility. Methods: This study utilised a genetic mouse model of obesity (ob/ob) to investigate obesity-induced male infertility. Control (C57BL/6) and obese (ob/ob) male mice were euthanized at 16 weeks of age. Epididymides were harvested to extract sperm to measure sperm concentration, motility (total, progressive and non-progressive), kinematics (straight line velocity (VSL), curvilinear velocity (VCL), amplitude of lateral head displacement (ALH) and linearity (LIN)), and morphology via Computer-Aided Sperm Analysis (CASA). Additionally, the testes were harvested and fixed in formalin or frozen for subsequent analyses. Formalin-fixed testes underwent haematoxylin and eosin (H&E), immunohistochemistry (IHC) and immunofluorescent (IF) staining to investigate testicular morphology and assessment of EV localisation, respectively. EV-specific biomarkers such as CD63, CD9 and ANXA1 were targeted. Frozen testes were utilised for EV extraction and further western blotting (WB) analyses. Spermatogenesis was investigated via WB for spermatogenesis-related proteins (IGF1, INSL6, and PRDX2). For EV extraction, frozen testes were dissociated and underwent differential centrifugation after which WB was performed to confirm EV isolation by targeting EV-specific biomarkers. Results: At 16-weeks old, obese mice had significantly increased body weights (P<0.0001) and fasting blood glucose levels (P=0.0004). Sperm concentration remained unchanged between groups. Obese spermatozoa had significantly reduced total (P<0.0001), progressive (P=0.009) and non-progressive (P<0.0001) motility when compared to controls. Kinematic measurements indicated reduced VSL (P=0.0064), VCL (P<0.0001) and ALH (P<0.0001) parameters for obese sperm, with LIN remaining unchanged. Obese mice presented with significantly increased abnormal spermatozoa (P<0.0001) and showed significant increased head and midpiece defects (P<0.0001 and P=0.0089, respectively). Obese testicular morphology displayed decreased seminiferous tubular area (P=0.0287) and diameter (P=0.0252), decreased lumen diameter (P=0.0011), as well as a high presence of vacuolation and disorganised cellular arrangements. IHC staining showed visible differences in EV localisation between control and obese testes for CD63, CD9 and ANXA1, with IF staining showing a decreased mean fluorescent intensity for CD63 (P=0.0345) in obese testes compared to controls. WB for IGF1, INSL6 and PRDX2 remained unchanged. WB for extracted EVs (CD63, CD9 and ANXA1) could not be quantified. Conclusion: Results from this study showed obesity-induced alterations in sperm parameters. Additionally, the testes and EV localisation results suggest a significant difference in CD63 EV marker compared to controls, suggesting less EVs in the ob/ob mice. Further investigation is required to delve deeper into the association between obesity, EV dynamics and testicular functions."]},{"key":"dc:title","label":"Title","values":["Obesity-Induced Alterations in Sperm Quality, Testicular Morphology, and Extracellular Vesicle Localisation in Male Mice"]}]}],"canonical_facts":{"dc:contributor.advisor":["Skosana, Bongekile Trisha","Marais, Erna"],"dc:contributor.other":["Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Division of Medical Physiology."],"dc:creator":["Uys, Sharnah"],"dc:date.accessioned":["2026-04-08T09:54:59Z"],"dc:date.available":["2026-04-08T09:54:59Z"],"dc:date.issued":["2026-03"],"dc:description":["Thesis (MSc)--Stellenbosch University, 2026.","Uys, S. 2026. Obesity-Induced Alterations in Sperm Quality, Testicular Morphology, and Extracellular Vesicle Localisation in Male Mice. Unpublished masters thesis. Stellenbosch: Stellenbosch University [online]. Available: https://scholar.sun.ac.za/items/cf57fd1c-cc36-4e58-b690-6f5273ad443a"],"dc:description.abstract":["Background: Infertility is a recognised reproductive health issue affecting individuals worldwide. An increase in male infertility has been reported, with concurrent evidence of declining semen quality. Obesity is a major contributor to infertility, with obese men presenting with decreased testosterone levels, impaired spermatogenesis and reduced sperm function. Extracellular vesicles (EVs), which facilitate cell-to-cell communication, have been discovered in various biological fluids. Specifically, in the male reproductive system, testicular-derived EVs have been reported to regulate testosterone production and spermatogenesis. There is a significant gap in knowledge regarding the effects of obesity on testicular EVs and their potential role in altering the testicular microenvironment. Therefore, examining testicular-derived EVs may provide an understanding of the association between obesity and male infertility. Methods: This study utilised a genetic mouse model of obesity (ob/ob) to investigate obesity-induced male infertility. Control (C57BL/6) and obese (ob/ob) male mice were euthanized at 16 weeks of age. Epididymides were harvested to extract sperm to measure sperm concentration, motility (total, progressive and non-progressive), kinematics (straight line velocity (VSL), curvilinear velocity (VCL), amplitude of lateral head displacement (ALH) and linearity (LIN)), and morphology via Computer-Aided Sperm Analysis (CASA). Additionally, the testes were harvested and fixed in formalin or frozen for subsequent analyses. Formalin-fixed testes underwent haematoxylin and eosin (H&E), immunohistochemistry (IHC) and immunofluorescent (IF) staining to investigate testicular morphology and assessment of EV localisation, respectively. EV-specific biomarkers such as CD63, CD9 and ANXA1 were targeted. Frozen testes were utilised for EV extraction and further western blotting (WB) analyses. Spermatogenesis was investigated via WB for spermatogenesis-related proteins (IGF1, INSL6, and PRDX2). For EV extraction, frozen testes were dissociated and underwent differential centrifugation after which WB was performed to confirm EV isolation by targeting EV-specific biomarkers. Results: At 16-weeks old, obese mice had significantly increased body weights (P<0.0001) and fasting blood glucose levels (P=0.0004). Sperm concentration remained unchanged between groups. Obese spermatozoa had significantly reduced total (P<0.0001), progressive (P=0.009) and non-progressive (P<0.0001) motility when compared to controls. Kinematic measurements indicated reduced VSL (P=0.0064), VCL (P<0.0001) and ALH (P<0.0001) parameters for obese sperm, with LIN remaining unchanged. Obese mice presented with significantly increased abnormal spermatozoa (P<0.0001) and showed significant increased head and midpiece defects (P<0.0001 and P=0.0089, respectively). Obese testicular morphology displayed decreased seminiferous tubular area (P=0.0287) and diameter (P=0.0252), decreased lumen diameter (P=0.0011), as well as a high presence of vacuolation and disorganised cellular arrangements. IHC staining showed visible differences in EV localisation between control and obese testes for CD63, CD9 and ANXA1, with IF staining showing a decreased mean fluorescent intensity for CD63 (P=0.0345) in obese testes compared to controls. WB for IGF1, INSL6 and PRDX2 remained unchanged. WB for extracted EVs (CD63, CD9 and ANXA1) could not be quantified. Conclusion: Results from this study showed obesity-induced alterations in sperm parameters. Additionally, the testes and EV localisation results suggest a significant difference in CD63 EV marker compared to controls, suggesting less EVs in the ob/ob mice. Further investigation is required to delve deeper into the association between obesity, EV dynamics and testicular functions."],"dc:identifier.uri":["https://scholar.sun.ac.za/handle/10019.1/135707"],"dc:language.iso":["en"],"dc:publisher":["Stellenbosch : Stellenbosch University"],"dc:title":["Obesity-Induced Alterations in Sperm Quality, Testicular Morphology, and Extracellular Vesicle Localisation in Male Mice"],"dc:type":["Thesis"]},"updated_at":"2026-07-24T04:40:12Z"}