The effect of calmodulin antagonists on scoliosis: bipedal C57BL/6 mice model

Abstract

C57BL6 mice are melatonin deficient from birth and have been shown to develop scoliosis when rendered bipedal. Our previous work suggested that tamoxifen and trifluoperozine may change the natural course of scoliosis in a chicken model. The objective of this study was to analyze whether the incidence of scoliosis or the magnitude of curves may be decreased by the administration of pharmacological agents tamoxifen or trifluoperozine in a mice scoliosis model. Sixty female 3-week-old C57BL6 mice underwent amputations of forelimbs and tails. Available 57 mice were divided into three groups, Group-I received no medications whereas Groups II and III received 10 mg TMX and 10 mg TMX + 10 mg TFP per liter of daily water supply, respectively. PA scoliosis X-rays were obtained at 20th and 40th weeks. Deformities were compared for incidence and the severity of the curves as well as disease progression or regression. At 20th week, overall, upper thoracic (UT), lower thoracic (T), and lumbar (L) scoliosis rates were similar (P = 0.531; P = 0.209; P = 0.926; P = 0.215, respectively) but thoraco-lumbar (TL) scoliosis rate was higher inTMX group (P = 0.036). However, at 40th week, although TL and L rates were similar (P = 0.628, P = 0.080), overall rate as well as the rates of UT and T scoliosis of TMX group were significantly lower (P = 0.001, P = 0.011, P = 0.001, respectively). As for curve magnitudes, T mean Cobb angle at 20th week was significantly higher in the C group (14 +/- 2.55) compared to TMX + TFP group (9 +/- 2.708; P = 0.033); at 40th week, TL mean Cobb angle was lower in the TMX + TFP group (17.50 +/- 3.45) compared to C (29.40 +/- 5.98; P = 0.031); and TMX group had lower TL Cobb angles compared to C (8.67 +/- 11.72) although not significant (P = 0.109). Double curve incidence at 40th week was significantly lower in TMX group compared to other groups (P = 0.001), triple curve incidence was lower in TMX + TFP and TMX groups, albeit not significant (P = 0.167). Between the 20th and 40th weeks, overall, double curve, and UT scoliosis rates showed an increase in C and TMX + TFP groups whereas TMX group showed a decline (P = 0.01, P = 0.002, P = 0.007, respectively). When specific regions were compared a similar significant difference was observed (P = 0.012 for upper thoracic; P = 0.018 for thoracic; P = 0.047 for thoraco-lumbar). This study has demonstrated that TMX is effective in changing the natural history of scoliotic deformities in C57BL6 mice model favorably.

Selective estrogen receptor modulation prevents scoliotic curve progression: radiologic and histomorphometric study on a bipedal C57Bl6 mice model

“…Therefore, AIS is modeled as a neuromuscular spinal deformity in which the melatonin effect on paraspinal muscle tonus may be by way of calmodulin antagonism, in that the absence of calmodulin antagonism in melatonin deficient scoliosis models may be the consequence of a paraspinal muscle tone imbalance eventually leading to scoliotic curves [17]. In previous studies, TMX as a calmodulin antagonist was used to replace the anticalmodulin effect of melatonin on melatonin deficient scoliotic mice and pinealectomized scoliotic chicken models [17,19]. Those results demonstrated that TMX did not have an effect on the incidence of scoliosis but decreased the curve progression and even reversed the curve magnitudes in some cases.…”

“…Eighty 3-week-old female C57BL6 (genetically altered to be melatonin deficient) mice were rendered bipedal by amputating the forelimbs at a high humeral level and tails at the root under general anesthesia as described previously [18,24] and divided into three groups: (1) TMX (n = 30); (2) RLX (n = 25); (3) Control (n = 25). In our previous experience, the death rate in the TMX group was relatively higher; therefore, we allocated more animals to the TMX group to compensate possible loss [19]. All animals received postoperative pain control and antibiotic prophylaxis and were kept in climate and photoperiodically controlled (12 h light, 12 h dark) standard mouse cages (5 per cage) with the food and water provided ad libidum from the top of the cage to encourage bipedal upright posture.…”

“…Dose of TMX and RLX were based of previous mice studies [19,22]. Weekly adjustments were made on the dosages of medications in accordance with the weight gain of the animals and the amount of water consumed daily per animal.…”

“…1) with animals lying on a custom-made jig to account for the shape of the thorax and thus to prevent animals from rolling over to the side. These time points were chosen to monitor the development and progression of scoliosis, in accordance with previous studies [19]. On the X-rays, curves in different locations in the spinal column were identified.…”

“…Research results demonstrated that tamoxifen (TMX), a Selective Estrogen Receptor Modulator (SERM) as well as a Calmodulin (CaM) antagonist, does not prevent the occurrence of the scoliotic deformities in either the pinealectomized chicken or melatonin deficient mice models; but decreases the rate of progression of deformity in both [17,19]. In addition, TMX may induce a reversal of the curves in significantly higher number of animals compared to controls.…”