Intracellular Ca2+ regulating proteins in vascular smooth muscle cells are altered with type 1 diabetes due to the direct effects of hyperglycemia.
BACKGROUND
Diminished calcium (Ca2+) transients in response to physiological agonists have been reported in vascular smooth muscle cells (VSMCs) from diabetic animals. However, the mechanism responsible was unclear.


RESULTS
VSMCs from autoimmune type 1 Diabetes Resistant Bio-Breeding (DR-BB) rats and streptozotocin-induced rats were examined for levels and distribution of inositol trisphosphate receptors (IP3R) and the SR Ca2+ pumps (SERCA 2 and 3). Generally, a decrease in IP3R levels and dramatic increase in ryanodine receptor (RyR) levels were noted in the aortic samples from diabetic animals. Redistribution of the specific IP3R subtypes was dependent on the rat model. SERCA 2 was redistributed to a peri-nuclear pattern that was more prominent in the DR-BB diabetic rat aorta than the STZ diabetic rat. The free intracellular Ca2+ in freshly dispersed VSMCs from control and diabetic animals was monitored using ratiometric Ca2+ sensitive fluorophores viewed by confocal microscopy. In control VSMCs, basal fluorescence levels were significantly higher in the nucleus relative to the cytoplasm, while in diabetic VSMCs they were essentially the same. Vasopressin induced a predictable increase in free intracellular Ca2+ in the VSMCs from control rats with a prolonged and significantly blunted response in the diabetic VSMCs. A slow rise in free intracellular Ca2+ in response to thapsigargin, a specific blocker of SERCA was seen in the control VSMCs but was significantly delayed and prolonged in cells from diabetic rats. To determine whether the changes were due to the direct effects of hyperglycemica, experiments were repeated using cultured rat aortic smooth muscle cells (A7r5) grown in hyperglycemic and control conditions. In general, they demonstrated the same changes in protein levels and distribution as well as the blunted Ca2+ responses to vasopressin and thapsigargin as noted in the cells from diabetic animals.


CONCLUSIONS
This work demonstrates that the previously-reported reduced Ca2+ signaling in VSMCs from diabetic animals is related to decreases and/or redistribution in the IP3R Ca2+ channels and SERCA proteins. These changes can be duplicated in culture with high glucose levels.
Searls
Yvonne M
YM
Department of Physical Therapy and Rehabilitation Science, University of Kansas Medical Center, Kansas City, KS 66160, USA.
Loganathan
Rajprasad
R
Smirnova
Irina V
IV
Stehno-Bittel
Lisa
L
D000818:Animals; D001011:Aorta / Q000378:metabolism; D001786:Blood Glucose / Q000378:metabolism; D001835:Body Weight; D002118:Calcium / Q000378:metabolism*; D003921:Diabetes Mellitus, Experimental / Q000097:blood / Q000503:physiopathology; D003922:Diabetes Mellitus, Type 1 / Q000097:blood / Q000150:complications* / Q000473:pathology / Q000503:physiopathology*; D006943:Hyperglycemia / Q000378:metabolism* / Q000473:pathology; D007150:Immunohistochemistry; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D051381:Rats; D011913:Rats, Inbred BB; D017207:Rats, Sprague-Dawley; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000378:metabolism
C506948:Atp2a2 protein, rat; C506951:Atp2a3 protein, rat; D001786:Blood Glucose; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D002118:Calcium