ABSTRACT
The secretion of insulin from pancreatic ß-cells is triggered by the influx of Ca(2+) through voltage-dependent Ca(2+) channels. The resulting elevation of intracellular calcium ([Ca(2+)](i)) triggers additional Ca(2+) release from internal stores. Less well understood are the mechanisms involved in Ca(2+) mobilization from internal stores after activation of Ca(2+) influx. The mobilization process is known as calcium-induced calcium release (CICR). In this study, our goal was to investigate the existence of and the role of caffeine-sensitive ryanodine receptors (RyRs) in a rat pancreatic ß-cell line, INS-1 cells. To measure cytosolic and stored Ca(2+), respectively, cultured INS-1 cells were loaded with fura-2/AM or furaptra/AM. [Ca(2+)](i) was repetitively increased by caffeine stimulation in normal Ca(2+) buffer. However, peak [Ca(2+)](i) was only observed after the first caffeine stimulation in Ca(2+) free buffer and this increase was markedly blocked by ruthenium red, a RyR blocker. KCl-induced elevations in [Ca(2+)](i) were reduced by pretreatment with ruthenium red, as well as by depletion of internal Ca(2+) stores using cyclopiazonic acid (CPA) or caffeine. Caffeine-induced Ca(2+) mobilization ceased after the internal stores were depleted by carbamylcholine (CCh) or CPA. In permeabilized INS-1 cells, Ca(2+) release from internal stores was activated by caffeine, Ca(2+), or ryanodine. Furthermore, ruthenium red completely blocked the CICR response in permeabilized cells. RyRs were widely distributed throughout the intracellular compartment of INS-1 cells. These results suggest that caffeine-sensitive RyRs exist and modulate the CICR response from internal stores in INS-1 pancreatic ß-cells.
ABSTRACT
Vascular smooth muscle cells can obtain a proliferative function in environments such as atherosclerosis in vivo or primary culture in vitro. Proliferation of vascular smooth muscle cells is accompanied by changes in ryanodine receptors (RyRs). In several studies, the cytosolic Ca(2+) response to caffeine is decreased during smooth muscle cell culture. Although caffeine is commonly used to investigate RyR function because it is difficult to measure Ca(2+) release from the sarcoplasmic reticulum (SR) directly, caffeine has additional off-target effects, including blocking inositol trisphosphate receptors and store-operated Ca(2+) entry. Using freshly dissociated rat aortic smooth muscle cells (RASMCs) and cultured RASMCs, we sought to provide direct evidence for the operation of RyRs through the Ca(2+)- induced Ca(2+)-release pathway by directly measuring Ca(2+) release from SR in permeabilized cells. An additional goal was to elucidate alterations of RyRs that occurred during culture. Perfusion of permeabilized, freshly dissociated RASMCs with Ca(2+) stimulated Ca(2+) release from the SR. Caffeine and ryanodine also induced Ca(2+) release from the SR in dissociated RASMCs. In contrast, ryanodine, caffeine and Ca(2+) failed to trigger Ca(2+) release in cultured RASMCs. These results are consistent with results obtained by immunocytochemistry, which showed that RyRs were expressed in dissociated RASMCs, but not in cultured RASMCs. This study is the first to demonstrate Ca(2+) release from the SR by cytosolic Ca(2+) elevation in vascular smooth muscle cells, and also supports previous studies on the alterations of RyRs in vascular smooth muscle cells associated with culture.
ABSTRACT
The aim of this study was to delineate the shape of the popliteus muscle and determine the correct motor point site for treating spasticity. A total of 22 legs from 13 fresh Korean cadavers were evaluated. The x-axis was set as a transverse line across the lateral and medial epicondyle of the femur and the y-axis as a vertical line at the midpoint of the medial malleolus of the tibia and lateral malleolus of the fibula. The popliteus muscle is an obtuse triangle in shape. Superior, medial, and inferior angles were 27.2±4.3°, 114.8±19.8°, and 38.0±18.8° respectively. The lengths of the superior, medial, and lateral sides of the triangle were 7.6±1.0 cm, 6.2±1.0 cm, and 11.9±1.5 cm respectively. Nerve branches ran superficially on the periosteum of the tibia and entered the popliteus on its superficial surface. The diverging point of the nerve branch entered the popliteus from the tibial nerve located at the midline of the popliteal fossa and 17% of the leg length above the intercondylar line. Most nerve entry points (83.3%) were within a 2.0×3.0 cm rectangle with the center located at -1.0 cm (-7%) on the x-axis and -3.3 cm (-9%) on the y-axis.
ABSTRACT
BACKGROUND: The estrogen receptor plays a critical role in breast cancer development and progression. So the genetic polymorphism of ER-alpha gene could affect cancer risk and phenotype. MATERIALS AND METHODS: We fully sequenced the ER-alpha gene to investigate its single nucleotide polymorphisms (SNPs) in 100 breast cancer patients and 100 controls. RESULTS: Six novel polymorphism in the control and 11 in cancer patients were found. The G/G genotype at C975 G and A/A genotype at G1782 A had a protective effect against breast cancer when compared to other genotypes (OR = 0.3 and 0.3, respectively). 975G allele was associated inversely with the p53 expression and positively with the bcl-2 expression in cancer with borderline significance. Combining this result with our previous study, these associations became more significant (p = 0.005 and 0.019, respectively). CONCLUSION: Polymorphisms in ER-alpha gene can affect the breast cancer susceptibility and may be related to other protein expression, such as p53 and bcl-2.
