The Top 5 Most Asked Questions On Z-VAD-FMK

1B). Both male and female CaR obKO mice and their heterozygous and control littermates were used for the experiments. Cre expression and activity in Col 3.6-Cre+/? transgenic mice are primarily detectable in calvarial and long bone preosteoblasts, osteoblasts, and osteocytes, and, to a much lesser degree, in kidney, liver, skin, testes, and ovaries.17 We confirmed the skeletal predominance of this expression pattern in the conditional knockouts (Fig. 1D) and noted low levels of expression of Cre in the lungs, heart, and kidneys. Necropsy of 5-day-old knockouts revealed immaturity of the lungs and kidneys (not shown), possibly resulting from the knockout of the CaR in the lung and renal fibroblasts. Nevertheless, there was no evidence of renal failure or substantial renal dysfunction in these mice by blood chemistries (Table 2). CaR obKO mice exhibited significantly reduced expression (by ?70%) of CaR transcripts in their humeral cortices, whereas heterozygotes expressed ?50% of the CaR transcripts compared with control mice (Fig. 1E). Residual expression of the CaR mRNA in bones from CaR obKOs may be attributed to the presence of nonosteoblastic cells in the tissue preparations that did not express Col 3.6-Cre, necessary for gene excision. Expression of the CaR protein in osteoblasts and osteocytes is absent, as shown by immunofluorescence studies of frozen sections of tibias from the CaR obKO animals (Fig. 1F). When primary antibodies were omitted, no immunofluorescence was visible (not shown). At birth, the weights of CaR obKO mice were significantly reduced (by 18%, p? with their littermates (Fig. 1G). From the second week of life, the CaR obKO mice suffered multiple fractures, particularly in the tibias, which impaired ambulation. CaR obKO mice died in either the third or fourth week of life, at which point their weights were ?70% less than their littermates. They lived longer in smaller litters, presumably because of reduced competition for nursing. There were no significant differences in serum levels of phosphorus, magnesium, sodium, chloride, creatinine, liver enzymes (alanine aminotransferase, aspartate aminotransferase), and total protein in knockouts and heterozygotes at 21 days of age compared with controls (Table 2). We noted mildly elevated serum total calcium in CaR obKO mice (12.5?��?0.35?mg/dL) compared with control (11.5?��?0.15?mg/dL) and heterozygous CaR obKO (11.4?��?0.22?mg/dL) littermates (p?