Concentration Of Calbindin Research Articles

La inmunorreactividad a Calbindina en células de purkinje del cerebelo de ratones no es afectada por la infección con virus de la rabia

Introducción: La calbindina (CB) es una proteína reguladora del calcio intracelular y la célula de Purkinje del cerebelo es la neurona con más alta concentración de CB. Se ha demostrado pérdida de inmunorreactividad a CB en diferentes áreas del sistema nervioso en ratones inoculados con virus de la rabia, pero faltaba estudiar este fenómeno en el cerebelo. Objetivo: Determinar el efecto de la inoculación con virus de la rabia sobre la expresión de CB en células de Purkinje del cerebelo de ratones. Metodología: Se inocularon ratones con el virus por vía intramuscular. Se sacrificaron los animales cuando alcanzaron la fase avanzada de la enfermedad y se fijaron mediante perfusión intracardiaca con paraformaldehído al 4%. Se les extrajo el cerebelo y se hicieron cortes sagitales de 50 micrómetros de espesor en un vibrátomo. Estos se procesaron mediante inmunohistoquímica para revelar la presencia de CB o de antígenos del virus de la rabia. El mismo procedimiento se realizó con animales no infectados (controles). Resultados: Las células de Purkinje fueron masivamente infectadas con el virus de la rabia. En las imágenes panorámicas observadas en el microscopio se comprobó que sólo estas células fueron inmunorreactivas a CB. No se hallaron diferencias significativas en la inmunorreactividad a CB, evaluada por densitometría óptica, entre los animales infectados y los controles. Conclusión: La expresión de CB en las células de Purkinje del cerebelo parece no afectarse por la infección con rabia, a diferencia de lo que se ha demostrado en otras áreas del sistema nervioso del ratón.

Cerebrospinal Fluid Calbindin D Concentration as a Biomarker of Cerebellar Disease Progression in Niemann-Pick Type C1 Disease.

Niemann-Pick type C (NPC) 1 disease is a rare, inherited, neurodegenerative disease. Clear evidence of the therapeutic efficacy of 2-hydroxypropyl-β-cyclodextrin (HPβCD) in animal models resulted in the initiation of a phase I/IIa clinical trial in 2013 and a phase IIb/III trial in 2015. With clinical trials ongoing, validation of a biomarker to track disease progression and serve as a supporting outcome measure of therapeutic efficacy has become compulsory. In this study, we evaluated calcium-binding protein calbindin D-28K (calbindin) concentrations in the cerebrospinal fluid (CSF) as a biomarker of NPC1 disease. In the naturally occurring feline model, CSF calbindin was significantly elevated at 3 weeks of age, prior to the onset of cerebellar dysfunction, and steadily increased to >10-fold over normal at end-stage disease. Biweekly intrathecal administration of HPβCD initiated prior to the onset of neurologic dysfunction completely normalized CSF calbindin in NPC1 cats at all time points analyzed when followed up to 78 weeks of age. Initiation of HPβCD after the onset of clinical signs (16 weeks of age) resulted in a delayed reduction of calbindin levels in the CSF. Evaluation of CSF from patients with NPC1 revealed that calbindin concentrations were significantly elevated compared with CSF samples collected from unaffected patients. Off-label treatment of patients with NPC1 with miglustat, an inhibitor of glycosphingolipid biosynthesis, significantly decreased CSF calbindin compared with pretreatment concentrations. These data suggest that the CSF calbindin concentration is a sensitive biomarker of NPC1 disease that could be instrumental as an outcome measure of therapeutic efficacy in ongoing clinical trials.

Estrogen and a calcium flux dependent factor modulate the calbindin gene expression in the uterus of laying hens

The dependency of calbindin 28K synthesis on estrogen and vitamin D and its relationship with calcium transfer were investigated in the uterus of laying hens by dot blot hybridization analysis using as a probe a cDNA coding for calbindin. Estrogen stimulated growth of the oviduct and uterine calbindin systhesis in juvenile D-deficient female chicks. In laying hens, calbindin mRNA increased most markedly during shell deposition but calbindin concentrations did not fluctuate during the ovulatory cycle. Suppression of shell formation within a few hours reduced calbindin mRNA levels and lowered uterine calbindin concentrations when egg expulsions were continued for several days. The concentration of calbindin and its mRNA increased when shell formation resumed in hens previously laying shell-less eggs. These increases were maintained in hens parathyroidectomized just before shell resumption. Lowering dietary calcium decreased uterine calcium transfer and calbindin concentration but its mRNA level was unaffected. It is suggested that uterine calbindin synthesis is regulated in a tissue-specific manner through transcriptional mechanisms irrespective of change in vitamin D; calbindin synthesis is stimulated by estrogens as part of its effect on oviductal growth but its regulation predominently involves a calcium flux dependent factor associated with shell calcification.

Regulation of calbindin D 28K and its mRNA in the intestine of the domestic hen

Intestinal calbindin synthesis in laying hens was analyzed to assess controlling factors operating during egg formation. In the absence of vitamin D, calbindin was not induced by estrogen and testosterone. In immature vitamin D-replete pullet, blood levels of 1,25(OH) 2D 3 increased in response to estrogen but the duodenal concentration of calbindin and its mRNA were increased only when testosterone was given together with estrogen. The plasma concentration of 1,25(OH) 2D 3 and the duodenal levels of calbindin and its mRNA were substantially higher in laying hens than in immature pullets. No differences in these parameters were observed between the stages of the ovulatory cycle. Suppression of shell formation for a week decreased the concentration of 1,25(OH) 2D 3 and of duodenal calbindin but did not affect the level of its mRNA. When egg shell formation resumed in hens previously laying shell-less eggs, the concentrations of 1,25(OH) 2D 3 and of calbindin and its mRNA increased toward the end of shell formation. A most important factor regulating intestinal calbindin synthesis in laying hens turned out to be 1,25(OH) 2D 3. Intestinal calbindin mRNA is more stable in laying hens than in young birds as its concentration declines more slowly when the stimulation provided by 1,25(OH) 2D 3 is withdrawn, as occurs following suppression of shell formation and after parathyroidectomy in laying birds. Intestinal calbindin mRNA is therefore increased by a process other than increasing 1,25(OH) 2D 3 formation. The factor influencing the stability of this mRNA in laying hens could be calcium. It is concluded that in hens the increased duodenal calbindin synthesis elicited by plasma 1,25(OH) 2D 3 at sexual maturity primarily involves a transcriptional process and the stabilization of the mRNA. Shell formation further increases the calbindin concentration, possibly by more effective translation of the calbindin mRNA.

Calcium-binding proteins are concentrated in the CA2 field of the monkey hippocampus: a possible key to this region's resistance to epileptic damage

Previous immunocytochemical studies have shown a heterogeneous distribution of parvalbumin (PA) and calbindin (CB) in the rat hippocampal formation. The results of the present study showed a heterogeneous distribution of PA and CB in primate Ammon's horn. The density and intensity of immunoreactivity for both of these calcium-binding proteins was greatest in CA2 as compared to CA1 and CA3. CB-immunoreactivity was localized to the cell bodies, dendrites, and axon initial segments of pyramidal cells whereas PA-immunostaining was found in the axon terminals, dendrites and cell bodies of interneurons that have features similar to GABAergic inhibitory neurons. Based on previous studies that have shown a protective role of calcium-binding proteins in neurons exposed to hyperstimulation, these results suggest that the resistance of CA2 pyramidal cells in temporal lobe epilepsy is due to the high concentration of CB and PA in this region of Ammon's horn.

Peptidyl-prolyl cis-trans isomerase does not affect the Pro-43 cis-trans isomerization rate in folded calbindin D 9k

The calcium-binding protein calbindin D 9k has previously been shown to exist in two folded forms only differing in the proline cis-trans isomerism of the Gly-42-Pro-43 amide bond. This bond is located in a flexible loop connecting the two EF-hand Ca 2+ sites. Calbindin D 9k therefore constitutes a unique test case for investigating if the recently discovered enzyme peptidyl-prolyl cis-trans isomerase (PPIase) can affect the cis-trans exchange rate in a folded protein. The 1H NMR saturation transfer technique has been used to measure the rate of interconversion between the cis and trans forms of calbindin in the presence of PPIase (PPIase:calbindin concentration ratio 1:10) at 35°C. No rate enhancement could be detected.

Differential regulation of calbindin-D28k mRNA in the intestine and eggshell gland of the laying hen

The effect of shell calcification and 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) on calbindin-D28K (previously known as vitamin D-dependent calcium-binding protein) and calbindin mRNA was investigated in the intestine and eggshell gland (ESG) of juvenile female chicks, laying hens and non-laying female birds with active gonads. Increasing amounts of 1,25-(OH)2D3 were fed to laying hens and juvenile birds treated with oestradiol to develop the ESG. The intestinal concentration of calbindin was increased 30-fold by 1,25-(OH)2D3 in chicks treated with oestradiol and fed a vitamin D-deficient diet. In these same animals, 1,25-(OH)2D3 had no effect on the formation of calbindin mRNA or calbindin in the ESG even though fully viable 1,25-(OH)2D3 receptors are present in this tissue. In laying birds fed adequate amounts of vitamin D3, intestinal, but not ESG, calbindin was increased by the addition of 1,25-(OH)2D3 to the diet. At the onset of egg production the concentrations of calbindin and calbindin mRNA were increased in the intestine and ESG. This increase occurred within the period of calcification of the first egg, through a process unaffected by vitamin D. Calcification of the first egg increased the concentration of calbindin in the ESG by eight- to tenfold, although the concentration of calbindin mRNA was increased by only two- to threefold. These results suggest that the induction of calbindin synthesis by 1,25-(OH)2D3 or by the egg calcification process is associated with an increase in the concentration of calbindin mRNA in the ESG and intestine.(ABSTRACT TRUNCATED AT 250 WORDS)

Increases in calbindin D 28K mRNA in the uterus of the domestic fowl induced by sexual maturity and shell formation

Uterine concentrations of calbindin D 28K mRNA were measured in immature pullets and laying hens by dot-blot hybridization using a [ 32P]cRNA probe prepared from the calbindin cDNA. In immature pullets, estrogen increased the calbindin mRNA level and the plasma concentration of 1,25-dihydroxyvitamin D 3 [1,25-(OH) 2D 3]. When testosterone was administered with estrogen there was a further increase in calbindin and its mRNA and an increase in the free 1,25-(OH) 2D 3 index calculated as the ratio of the molar concentrations of total 1,25-(OH) 2D 3 and vitamin D-binding protein (DBP). In laying hens the uterine concentration of calbindin mRNA was low 4 hr after ovulation, but increased most markedly 12 and 18 hr later, when shell calcification took place. Calbindin concentration remained unchanged during the different stages of egg formation but was much higher in laying hens than in pullets treated with sex steroids. Suppression of shell formation by premature expulsion of the egg decreased the concentrations of calbindin mRNA and uterine calbindin and the free 1,25-(OH) 2D 3 index in the plasma. A concomitant increase in calbindin and its mRNA was observed at resumption of shell formation in hens previously laying shell-less eggs. With-drawal of food for 44 hr decreased the uterine concentration of calbindin and its mRNA without a change in the free 1,25-(OH) 2D 3 index in the blood. It is concluded that the synthesis of uterine calbindin is stimulated primarily at sexual maturity and at calcification of the first shell by transcriptional processes. The daily increase in calbindin mRNA associated with shell formation and the absence of a concomitant change in calbindin concentration suggest that post-transcriptional processes exist and that stimuli other than the sex steroid or the 1,25-(OH) 2D 3 are involved in regulation of calbindin synthesis in the uterus.

Tissue-specific regulation of the concentration of Calbindin D 28K mRNA in the developing chicken

A BamHI-HindIII restriction fragment containing the 5′-terminal portion of the gene encoding chicken Calbindin D 28K was sequenced and used as a probe in Northern-blot hydridizations to RNA extracted from the brain and intestine of chickens at various stages of development. In both tissues Calbindin D 28K mRNA consists of a family of three species, which differ by size. In the intestine Calbindin D 28K mRNAs appear at hatching and reach a peak at day 7. In the brain the same RNA species are easily detected at least 7 days before hatching, show a moderate increase at hatching and remain essentially constant during the first 10 days of adult life. The concentration of Calbindin D 28K mRNAs in the intestine is strictly dependent on Vitamin D, while it is not in the cerebellum.

Egg Shell Quality and Cholecalciferol Metabolism in Aged Laying Hens

Calcium-binding protein D28K (calbindin) synthesis, vitamin D metabolism and shell quality were investigated in young and aged laying hens fed diets containing either cholecalciferol (CC) or its 1-hydroxylated derivatives. Duodenal calbindin concentration was similar in the young and in the aged laying hens. Exogenous 1-hydroxylated CC derivatives increased duodenal calbindin concentration, regardless of age. Shell weight and shell density (mg/cm2) were significantly lower (P less than 0.01) in the aged than in the young hens. Egg shell weight and density tended to decrease along the clutch. The rate of decline was higher in aged than in young hens. Feeding aged hens a diet containing 5 micrograms 1,25-dihydroxycholecalciferol [1,25(OH)2CC] or 1 alpha-hydroxycholecalciferol per kilogram improved shell quality, slowed down the progressive reduction in shell quality during the clutch and increased culling and mortality. The results indicate a) that the capacity for expression of 1,25(OH)2D3 in the intestine is not altered by age and b) that prolonged feeding of 1-hydroxylated derivatives of vitamin D3 improves shell quality in aged laying hens and increases culling and mortality.

Relationship between the levels of calbindin synthesis and calbindin mRNA in chick intestine. Quantitation of calbindin mRNA.

An RNA-excess filter hybridization assay was established to measure the absolute amount of calbindin mRNA in chick tissues. The tissue with the highest level of mRNA is intestine, followed by kidney and cerebellum; the mRNA was not detected in liver and skin. Calbindin mRNA in intestine and kidney is vitamin D-dependent. The maximum concentration of calbindin and its mRNA found after dosing vitamin D-deficient chicks with dihydroxyvitamin D3 (1,25-(OH)2D3) is less than 5% of that found with vitamin D dosing. Secondary 1,25-(OH)2D3 stimulation produced greatly increased amounts of both calbindin mRNA and the protein, at least reaching levels similar to those found after vitamin D dosing. In this last case, each mucosal cell contains about 2000 calbindin mRNA molecules which are translated at a rate sufficient to account for the levels of calbindin found. Calbindin mRNA is translated most rapidly in the very short time periods after its release into the cytoplasm. 1,25-(OH)2D3 has two effects on calbindin mRNA formation: first, to permit the expression of the calbindin gene and a second effect, of slower onset but more persistent, which increases either the rate of calbindin gene transcription or the stability of calbindin mRNA.