Regional investigation of UT-B urea transporters in the rat brain
Recent studies have reported increased levels of urea in the aging brain and various neurological disorders. Additionally, these diseased tissues also have increased expression of the UT-B transporter that regulates urea transport in the brain. However, little is known regarding the actual UT-B prot...
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| Format: | Article |
| Language: | English |
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Elsevier
2023-12-01
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| Series: | Biochemistry and Biophysics Reports |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2405580823001449 |
| _version_ | 1797422558807588864 |
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| author | Farhana Pinki Derek A. Costello Gavin Stewart |
| author_facet | Farhana Pinki Derek A. Costello Gavin Stewart |
| author_sort | Farhana Pinki |
| collection | DOAJ |
| description | Recent studies have reported increased levels of urea in the aging brain and various neurological disorders. Additionally, these diseased tissues also have increased expression of the UT-B transporter that regulates urea transport in the brain. However, little is known regarding the actual UT-B protein distribution across the brain in either normal or diseased states. This current study investigated UT-B protein abundance across three regions of the rat brain – anterior, posterior and cerebellum. Endpoint RT-PCR experiments showed that there were no regional differences in UT-B RNA expression (NS, N = 3, ANOVA), whilst Western blotting confirmed no difference in the abundance of a 35 kDa UT-B protein (NS, N = 3–4, ANOVA). In contrast, there was a significant variation in a non-UT-B 100 kDa protein (P < 0.001, N = 3–4, ANOVA), which was also detected by anti-UT-B antibodies. Using the C6 rat astrocyte cell line, Western blot analysis showed that 48-h incubation in either 5 mM or 10 mM significantly increased a 30–45 kDa UT-B protein signal (P < 0.05, N = 3, ANOVA). Furthermore, investigation of compartmentalized C6 protein samples showed the 30–45 kDa signal in the membrane fraction, whilst the 100 kDa non-UT-B signal was predominantly in the cytosolic fraction. Finally, immunolocalization studies gave surprisingly weak detection of rat UT-B, except for strong staining of red blood cells in the cerebellum. In conclusion, this study confirmed that RNA expression and protein abundance of UT-B were equal across all regions of the rat brain, suggesting that urea levels were also similar. However, it also highlighted some of the technical challenges of studying urea transporters at the protein level. |
| first_indexed | 2024-03-09T07:34:02Z |
| format | Article |
| id | doaj.art-315dc0b0b6ad4e83b49b8b783b1058ef |
| institution | Directory Open Access Journal |
| issn | 2405-5808 |
| language | English |
| last_indexed | 2024-03-09T07:34:02Z |
| publishDate | 2023-12-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Biochemistry and Biophysics Reports |
| spelling | doaj.art-315dc0b0b6ad4e83b49b8b783b1058ef2023-12-03T05:41:55ZengElsevierBiochemistry and Biophysics Reports2405-58082023-12-0136101563Regional investigation of UT-B urea transporters in the rat brainFarhana Pinki0Derek A. Costello1Gavin Stewart2UCD School of Biology and Environmental Science, University College Dublin, Dublin 4, IrelandUCD School of Biomolecular and Biomedical Science, University College Dublin, Dublin 4, Ireland; UCD Conway Institute, University College Dublin, Dublin 4, IrelandUCD School of Biology and Environmental Science, University College Dublin, Dublin 4, Ireland; Corresponding author.Recent studies have reported increased levels of urea in the aging brain and various neurological disorders. Additionally, these diseased tissues also have increased expression of the UT-B transporter that regulates urea transport in the brain. However, little is known regarding the actual UT-B protein distribution across the brain in either normal or diseased states. This current study investigated UT-B protein abundance across three regions of the rat brain – anterior, posterior and cerebellum. Endpoint RT-PCR experiments showed that there were no regional differences in UT-B RNA expression (NS, N = 3, ANOVA), whilst Western blotting confirmed no difference in the abundance of a 35 kDa UT-B protein (NS, N = 3–4, ANOVA). In contrast, there was a significant variation in a non-UT-B 100 kDa protein (P < 0.001, N = 3–4, ANOVA), which was also detected by anti-UT-B antibodies. Using the C6 rat astrocyte cell line, Western blot analysis showed that 48-h incubation in either 5 mM or 10 mM significantly increased a 30–45 kDa UT-B protein signal (P < 0.05, N = 3, ANOVA). Furthermore, investigation of compartmentalized C6 protein samples showed the 30–45 kDa signal in the membrane fraction, whilst the 100 kDa non-UT-B signal was predominantly in the cytosolic fraction. Finally, immunolocalization studies gave surprisingly weak detection of rat UT-B, except for strong staining of red blood cells in the cerebellum. In conclusion, this study confirmed that RNA expression and protein abundance of UT-B were equal across all regions of the rat brain, suggesting that urea levels were also similar. However, it also highlighted some of the technical challenges of studying urea transporters at the protein level.http://www.sciencedirect.com/science/article/pii/S2405580823001449UreaUT-B transporterBrainRegionProtein |
| spellingShingle | Farhana Pinki Derek A. Costello Gavin Stewart Regional investigation of UT-B urea transporters in the rat brain Biochemistry and Biophysics Reports Urea UT-B transporter Brain Region Protein |
| title | Regional investigation of UT-B urea transporters in the rat brain |
| title_full | Regional investigation of UT-B urea transporters in the rat brain |
| title_fullStr | Regional investigation of UT-B urea transporters in the rat brain |
| title_full_unstemmed | Regional investigation of UT-B urea transporters in the rat brain |
| title_short | Regional investigation of UT-B urea transporters in the rat brain |
| title_sort | regional investigation of ut b urea transporters in the rat brain |
| topic | Urea UT-B transporter Brain Region Protein |
| url | http://www.sciencedirect.com/science/article/pii/S2405580823001449 |
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