• MeSH
• fyziologie buňky MeSH
• konfokální mikroskopie metody   MeSH
• pohyb MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH

• MeSH
• buněčná membrána ultrastruktura   MeSH
• cytoplazma ultrastruktura   MeSH
• experimentální sarkom ultrastruktura   MeSH
• konfokální mikroskopie MeSH
• organely ultrastruktura   MeSH
• videomikroskopie MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH

• MeSH
• audiovizuální záznam MeSH
• buněčné dělení MeSH
• krysa rodu rattus MeSH
• mikrofotografie MeSH
• nádorové buňky kultivované MeSH
• pohyb buněk MeSH
• sarkom ultrastruktura   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• zvířata MeSH

• MeSH
• konfokální mikroskopie přístrojové vybavení   MeSH
• mikrofotografie metody   trendy   MeSH
• věda MeSH

• MeSH
• experimentální sarkom MeSH
• fibroblasty fyziologie   chemie   MeSH
• finanční podpora výzkumu jako téma MeSH
• kalibrace MeSH
• krysa rodu rattus MeSH
• kultivované buňky MeSH
• listy rostlin cytologie   fyziologie   MeSH
• pohyb buněk MeSH
• zobrazování trojrozměrné MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• zvířata MeSH

We visualized insulin uptake in vivo across the apical membrane of the rat proximal tubule (PT) by confocal microscopy; we compared it with in vitro findings in a rat PT cell line (WKPT) using fluorescence microscopy and flow cytometry. Surface tubules were observed in vivo with a 633-nm single laser-illuminated real-time video-rate confocal scanning microscope in upright configuration for optical sectioning below the renal capsule. Fields were selected containing proximal and distal tubules; Cy5-labeled insulin was injected twice (the second time after approximately 140 min) into the right jugular vein, and the fluorescence signal (at 650-670 nm) was recorded. Fluorescence was detected almost immediately at the brush-border membrane (BBM) of PT cells only, moving inside cells within 30-40 min. As a measure of insulin uptake, the ratio of the fluorescence signal after the second injection to the first doubled (ratio: 2.11 +/- 0.26, mean +/- SE, n = 10), indicating a "priming," or stimulating, effect of insulin on its uptake mechanism at the BBM. This effect did not occur after pretreatment with intravenous lysine (ratio: 1.03 +/- 0.07, n = 6; P < 0.01). Cy2- or Cy3-labeled insulin uptake in a PT cell line in vitro was monitored by 488-nm excitation fluorescence microscopy using an inverted microscope. Insulin localized toward the apical membrane of these cells. Semiquantitative analysis of insulin uptake by flow cytometry also demonstrated a priming effect (upregulation) on insulin internalization in the presence of increasing amounts of insulin, as was observed in vivo; moreover, this effect was not seen with, or affected by, the similarly endocytosed ligand beta2-glycoprotein.

• MeSH
• buněčné linie MeSH
• distální tubuly ledvin cytologie   metabolismus   MeSH
• elektronová mikroskopie MeSH
• endocytóza fyziologie   MeSH
• epitelové buňky cytologie   metabolismus   ultrastruktura   MeSH
• fluorescenční mikroskopie metody   MeSH
• hypoglykemika farmakokinetika   MeSH
• inzulin farmakokinetika   MeSH
• konfokální mikroskopie metody   MeSH
• krysa rodu rattus MeSH
• potkani inbrední WKY MeSH
• potkani Sprague-Dawley MeSH
• proximální tubuly ledvin cytologie   metabolismus   MeSH
• průtoková cytometrie MeSH
• techniky in vitro MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• práce podpořená grantem MeSH

A new genetic disorder has been identified that results from mutation of THRA, encoding thyroid hormone receptor α1 (TRα1). Affected children have a high serum T3:T4 ratio and variable degrees of intellectual deficit and constipation but exhibit a consistently severe skeletal dysplasia. In an attempt to improve developmental delay and alleviate symptoms of hypothyroidism, patients are receiving varying doses and durations of T4 treatment, but responses have been inconsistent so far. Thra1(PV/+) mice express a similar potent dominant-negative mutant TRα1 to affected individuals, and thus represent an excellent disease model. We hypothesized that Thra1(PV/+) mice could be used to predict the skeletal outcome of human THRA mutations and determine whether prolonged treatment with a supraphysiological dose of T4 ameliorates the skeletal abnormalities. Adult female Thra1(PV/+) mice had short stature, grossly abnormal bone morphology but normal bone strength despite high bone mass. Although T4 treatment suppressed TSH secretion, it had no effect on skeletal maturation, linear growth, or bone mineralization, thus demonstrating profound tissue resistance to thyroid hormone. Despite this, prolonged T4 treatment abnormally increased bone stiffness and strength, suggesting the potential for detrimental consequences in the long term. Our studies establish that TRα1 has an essential role in the developing and adult skeleton and predict that patients with different THRA mutations will display variable responses to T4 treatment, which depend on the severity of the causative mutation.

• MeSH
• fyziologická kalcifikace MeSH
• kostní denzita MeSH
• lidé MeSH
• mutace MeSH
• myši inbrední C57BL MeSH
• myši transgenní MeSH
• myši MeSH
• thyroxin metabolismus   MeSH
• tyreoidální hormony, receptory alfa genetika   metabolismus   MeSH
• velikost těla MeSH
• vývojové onemocnění kostí genetika   metabolismus   patofyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Intramural MeSH