No Source for ORAI1 Gene Knockout Comfort Control Found

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Subsequently, their ORAI1 gene was cloned and sequenced to confirm the successful deletion of the start codon. After clone expansion, effective KO generation was again validated by PCR (Figure 2C) and confirmed by immunoblot analysis (Figure 2D). ... Experimental design of the strategy used to create total knockout of ORAI1 protein (ORAI1 KO).

Furthermore, current evidence supports that abnormal Orai1 expression or function underlies several disorders. Orai1 is, together with STIM1, the key element of SOCE, conducting the Ca2+ release-activated Ca2+ (CRAC) current and, in association with TRPC1, the store-operated Ca2+ (SOC) current.

(C) Maximum rate of EDL force production and relaxation for twitch (left) and tetanic (right) stimulation (∆F(mN/mm2)/∆t(second)). (D) Peak EDL-specific force during repetitive, high-frequency stimulation (60 stimulus trains, 500 ms duration trains, 50 Hz stimulation frequency, delivered every 2.5 s). Data are shown as means ± SEM. * p < 0.05 and ** p < 0.01 as evaluated by unpaired Student’s t-test, n = 12 muscles from seven control mice (three males and four females) and n = 9 muscles from five cOrai1 KO mice (three males and two females).

The difference in SOCE between ... was significant (p = 0.007). ... ORAI1 knockout (KO) leads to the downregulation of ORAI2 and ORAI3 messenger RNA (mRNA) expression....

Number of mice used: WT (n = 7), mdx (n = 7), mdx-Orai1 KO (n = 5), and Orai1 KO (n = 6). ... Eccentric contraction–induced damage in EDL muscles in the absence or presence of BTP2. … ... Eccentric contraction–induced damage in EDL muscles in the absence or presence of BTP2. (A and B) Specific force drop during 10 consecutive eccentric contractions in EDL muscles from WT (A, n = 4 muscles) and mdx (B, n = 4 muscles) mice while perfused in either control Ringer’s solution (black curve) or a Ringer’s solution supplemented with 10 µM BTP2 (red curves).

To knock out Orai1 expression, we used two approaches: (i) For the in vitro mechanistic studies, we isolated primary spinal cord microglia from Orai1fl/fl Cx3CR1-Cre/ERT2 mice or WT controls from postnatal day 0 (P0) to P2 stage, cultured them for 10 days, and added tamoxifen or vehicle to the culture medium for Orai1 deletion (see Materials and Methods).