{"id":{"repo_id":"buffalo","oai_identifier":"oai:ubir.buffalo.edu:10477/86824"},"canonical_url":"https://search.dev.ndltd.org/etd/buffalo/oai:ubir.buffalo.edu:10477/86824","repository":{"repo_id":"buffalo","name":"Buffalo","base_url":"https://ubir.buffalo.edu/oai/request"},"display":{"title":"The Mechanism of Cadmium Removal from Renal Cell Model Mediated by GMDTC","abstract":"M.S.","abstract_html":"M.S.","abstract_has_math":false,"creators":["Xiao, Bo"],"institution":"State University of New York at Buffalo","degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":null,"school":null,"contributors":["Ren, Xuefeng","Pharmacology and Toxicology"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2025,"date_issued":"2025-02-25T23:23:09Z","date_published":"2025-02-25T23:23:09Z","updated_at":"2026-07-27T19:05:37Z","subjects":["pharmacology","toxicology"],"languages":["eng"],"rights":["Users of works found in University at Buffalo Institutional Repository (UBIR) are responsible for identifying and contacting the copyright owner for permission to reuse. University at Buffalo Libraries do not manage rights for copyright-protected works and cannot assist with permissions.","Copyright retained by author."],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"http://hdl.handle.net/10477/86824","outbound_label":"Handle","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Ren, Xuefeng","Pharmacology and Toxicology"]},{"key":"dc:creator","label":"Author","values":["Xiao, Bo"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2025-02-25T23:23:09Z","2020","2020-07-29 14:59:06"]},{"key":"dc:publisher","label":"Institution","values":["State University of New York at Buffalo"]},{"key":"dc:type","label":"Dc Type","values":["Text","Thesis"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["pharmacology","toxicology"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["eng"]},{"key":"dc:rights","label":"Dc Rights","values":["Users of works found in University at Buffalo Institutional Repository (UBIR) are responsible for identifying and contacting the copyright owner for permission to reuse. University at Buffalo Libraries do not manage rights for copyright-protected works and cannot assist with permissions.","Copyright retained by author."]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["http://hdl.handle.net/10477/86824"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["M.S.","Cadmium (Cd) is classified as a known human carcinogen (IA Class) by the International Agency for Research on Cancer (IARC). Cadmium combines with metallothionein to form a complex which is transferred through blood circulation to the kidneys. Such complexes degrade in lysosomes and release free cadmium ions which cause the loss of calcium and vitamin D, leading to osteoporosis, diabetes, reproductive system problem, and other severe diseases. However, the body lacks the effective mechanisms to clear cadmium ions. Currently, the use of common chelating agents, e.g., EDTA, BAL, has proven disappointing due to their severe side effects and/or ineffectiveness in removing cadmium from the kidney. GMDTC, a newly synthesized DTC (dithiocarbamate) derivative, has been shown to be safe (very low toxicity) and efficient in removing cadmium and other metals from the kidneys of the mouse, rat, and rabbit, as well as from human proximal tubule cells (HK-2). The objective of this project is to clarify the mechanism of cadmium excretion mediated by GMDTC utilizing CRISPR/Cas9 technology to construct SGLT2, GLUT2 single and double-gene knockout (SGLT2-/-, GLUT2-/-, SGLT2-/-/GLUT2-/-) in HK-2 cell lines. These constructed models were used to investigate GMDTC and its potential to reduce cadmium-induced renal toxicity in vitro. The ability of GMDTC to react with cadmium to form a GMDTC-cadmium complex was shown through UV-VIS spectroscopy. It was observed that co-treatment of cadmium with GMDTC could increase cell viability in HK-2 cell line and decrease cadmium-induced toxicity. Cell viability also increased in cells treated with cadmium and GMDTC in GMDTC rescue experiments, indicating that GMDTC could attenuate cadmium-induced toxicity caused by intracellular cadmium accumulation. We hypothesize that the mechanism of cadmium removal from the kidney by GMDTC is through the glucose reabsorption pathway, consisting of SGLT2 and GLUT2 glucose transporters. We developed the CRISPR/Cas9 mediated SGLT2, GLUT2 single and double gene knockout cell lines from HK-2 cells. This served to determine the role of glucose reabsorption pathway in GMDTC’s ability of removing cadmium from kidney cells. Validation of successful knockouts was conducted using flow cytometer, western blot analysis, Sanger sequencing analysis, and Tracking of Insertions and Deletions by decomposition (TIDE) analysis. Two monoclonal cell lines for the single gene knockout and one for the double gene knockout were used for later experiments. Compared to control, cell viability decreased in both single and double gene knockout cell models in GMDTC rescue experiments, which indicates that GMDTC may not enter into cells and/or the GMDTC-cadmium complex cannot discharge from cells after SGLT2 and GLUT2 gene knockouts. Removal of cadmium from cells by GMDTC may rely on the assistance of these two glucose transporters. To verify this mechanism, two chemical inhibitors of SGLT2 and GLUT2 were used to assess the effects of GMDTC in reducing cadmium-induced toxicity in HK-2 cells. Cell viability significantly decreased with inhibition of either SGLT2 or GLUT2 protein carrier, suggesting that as a result of the inhibition of glucose transporters, GMDTC could not get into cells to form the GMDTC-cadmium complex which then would be excreted from cells. In summary, these results indirectly indicate that GMDTC protects against cadmium-induced toxicity by preventing the accumulation of cadmium in renal cells and by removing intracellular cadmium via the glucose reabsorption pathway. In conclusion, the results of this study suggest that GMDTC reduces cadmium-induced renal toxicity by preventing the accumulation and assisting in the removal of intracellular cadmium. The potential mechanism of GMDTC in removing cadmium in vitro is likely mediated by the glucose transporters, SGLT2 and GLUT2. Therefore, GMDTC may be clinically useful for the prevention of cadmium poisoning.","**To request an accessible version of the file(s) associated with this item, contact library@buffalo.edu. Please include the item's persistent URL [http://hdl.handle.net/. . .] in your request.**"]},{"key":"dc:format","label":"Dc Format","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["The Mechanism of Cadmium Removal from Renal Cell Model Mediated by GMDTC"]}]}],"canonical_facts":{"dc:contributor":["Ren, Xuefeng","Pharmacology and Toxicology"],"dc:creator":["Xiao, Bo"],"dc:date":["2025-02-25T23:23:09Z","2020","2020-07-29 14:59:06"],"dc:description":["M.S.","Cadmium (Cd) is classified as a known human carcinogen (IA Class) by the International Agency for Research on Cancer (IARC). Cadmium combines with metallothionein to form a complex which is transferred through blood circulation to the kidneys. Such complexes degrade in lysosomes and release free cadmium ions which cause the loss of calcium and vitamin D, leading to osteoporosis, diabetes, reproductive system problem, and other severe diseases. However, the body lacks the effective mechanisms to clear cadmium ions. Currently, the use of common chelating agents, e.g., EDTA, BAL, has proven disappointing due to their severe side effects and/or ineffectiveness in removing cadmium from the kidney. GMDTC, a newly synthesized DTC (dithiocarbamate) derivative, has been shown to be safe (very low toxicity) and efficient in removing cadmium and other metals from the kidneys of the mouse, rat, and rabbit, as well as from human proximal tubule cells (HK-2). The objective of this project is to clarify the mechanism of cadmium excretion mediated by GMDTC utilizing CRISPR/Cas9 technology to construct SGLT2, GLUT2 single and double-gene knockout (SGLT2-/-, GLUT2-/-, SGLT2-/-/GLUT2-/-) in HK-2 cell lines. These constructed models were used to investigate GMDTC and its potential to reduce cadmium-induced renal toxicity in vitro. The ability of GMDTC to react with cadmium to form a GMDTC-cadmium complex was shown through UV-VIS spectroscopy. It was observed that co-treatment of cadmium with GMDTC could increase cell viability in HK-2 cell line and decrease cadmium-induced toxicity. Cell viability also increased in cells treated with cadmium and GMDTC in GMDTC rescue experiments, indicating that GMDTC could attenuate cadmium-induced toxicity caused by intracellular cadmium accumulation. We hypothesize that the mechanism of cadmium removal from the kidney by GMDTC is through the glucose reabsorption pathway, consisting of SGLT2 and GLUT2 glucose transporters. We developed the CRISPR/Cas9 mediated SGLT2, GLUT2 single and double gene knockout cell lines from HK-2 cells. This served to determine the role of glucose reabsorption pathway in GMDTC’s ability of removing cadmium from kidney cells. Validation of successful knockouts was conducted using flow cytometer, western blot analysis, Sanger sequencing analysis, and Tracking of Insertions and Deletions by decomposition (TIDE) analysis. Two monoclonal cell lines for the single gene knockout and one for the double gene knockout were used for later experiments. Compared to control, cell viability decreased in both single and double gene knockout cell models in GMDTC rescue experiments, which indicates that GMDTC may not enter into cells and/or the GMDTC-cadmium complex cannot discharge from cells after SGLT2 and GLUT2 gene knockouts. Removal of cadmium from cells by GMDTC may rely on the assistance of these two glucose transporters. To verify this mechanism, two chemical inhibitors of SGLT2 and GLUT2 were used to assess the effects of GMDTC in reducing cadmium-induced toxicity in HK-2 cells. Cell viability significantly decreased with inhibition of either SGLT2 or GLUT2 protein carrier, suggesting that as a result of the inhibition of glucose transporters, GMDTC could not get into cells to form the GMDTC-cadmium complex which then would be excreted from cells. In summary, these results indirectly indicate that GMDTC protects against cadmium-induced toxicity by preventing the accumulation of cadmium in renal cells and by removing intracellular cadmium via the glucose reabsorption pathway. In conclusion, the results of this study suggest that GMDTC reduces cadmium-induced renal toxicity by preventing the accumulation and assisting in the removal of intracellular cadmium. The potential mechanism of GMDTC in removing cadmium in vitro is likely mediated by the glucose transporters, SGLT2 and GLUT2. Therefore, GMDTC may be clinically useful for the prevention of cadmium poisoning.","**To request an accessible version of the file(s) associated with this item, contact library@buffalo.edu. Please include the item's persistent URL [http://hdl.handle.net/. . .] in your request.**"],"dc:format":["application/pdf"],"dc:identifier":["http://hdl.handle.net/10477/86824"],"dc:language":["eng"],"dc:publisher":["State University of New York at Buffalo"],"dc:rights":["Users of works found in University at Buffalo Institutional Repository (UBIR) are responsible for identifying and contacting the copyright owner for permission to reuse. University at Buffalo Libraries do not manage rights for copyright-protected works and cannot assist with permissions.","Copyright retained by author."],"dc:subject":["pharmacology","toxicology"],"dc:title":["The Mechanism of Cadmium Removal from Renal Cell Model Mediated by GMDTC"],"dc:type":["Text","Thesis"]},"updated_at":"2026-07-27T19:05:37Z"}