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Different effects of valproic acid on photoreceptor loss in Rd1 and Rd10 retinal degeneration mice.

Mitton KP, Guzman AE, Deshpande M, Byrd D, DeLooff C, Mkoyan K, Zlojutro P, Wallace A, Metcalf B, Laux K, Sotzen J, Tran T - Mol. Vis. (2014)

Bottom Line: Nrl gene expression was decreased by 50%, while Crx gene expression was not affected.Rod-specific expression of Mef2c and Nr2e3 was decreased substantially by VPA treatment, while Rhodopsin and Pde6b gene expression was normal at P28.While daily treatment with VPA could significantly reduce photoreceptor loss in the rd1 model, VPA treatment slightly accelerated photoreceptor loss in the rd10 model.

View Article: PubMed Central - PubMed

Affiliation: Control of Gene Expression Laboratory and Pediatric Retinal Research Laboratory, Eye Research Institute, Oakland University, Rochester, MI.

ABSTRACT

Purpose: The histone-deacetylase inhibitor activity of valproic acid (VPA) was discovered after VPA's adoption as an anticonvulsant. This generated speculation for VPA's potential to increase the expression of neuroprotective genes. Clinical trials for retinitis pigmentosa (RP) are currently active, testing VPA's potential to reduce photoreceptor loss; however, we lack information regarding the effects of VPA on available mammalian models of retinal degeneration, nor do we know if retinal gene expression is perturbed by VPA in a predictable way. Thus, we examined the effects of systemic VPA on neurotrophic factor and Nrl-related gene expression in the mouse retina and compared VPA's effects on the rate of photoreceptor loss in two strains of mice, Pde6b(rd1/rd1) and Pde6b(rd10/rd10) .

Methods: The expression of Bdnf, Gdnf, Cntf, and Fgf2 was measured by quantitative PCR after single and multiple doses of VPA (intraperitoneal) in wild-type and Pde6b(rd1/rd1) mice. Pde6b(rd1/rd1) mice were treated with daily doses of VPA during the period of rapid photoreceptor loss. Pde6b(rd10/rd10) mice were also treated with systemic VPA to compare in a partial loss-of-function model. Retinal morphology was assessed by virtual microscopy or spectral-domain optical coherence tomography (SD-OCT). Full-field and focal electroretinography (ERG) analysis were employed with Pde6b(rd10/rd10) mice to measure retinal function.

Results: In wild-type postnatal mice, a single VPA dose increased the expression of Bdnf and Gdnf in the neural retina after 18 h, while the expression of Cntf was reduced by 70%. Daily dosing of wild-type mice from postnatal day P17 to P28 resulted in smaller increases in Bdnf and Gdnf expression, normal Cntf expression, and reduced Fgf2 expression (25%). Nrl gene expression was decreased by 50%, while Crx gene expression was not affected. Rod-specific expression of Mef2c and Nr2e3 was decreased substantially by VPA treatment, while Rhodopsin and Pde6b gene expression was normal at P28. Daily injections with VPA (P9-P21) dramatically slowed the loss of rod photoreceptors in Pde6b(rd1/rd1) mice. At age P21, VPA-treated mice had several extra rows of rod photoreceptor nuclei compared to PBS-injected littermates. Dosing started later (P14) or dosing every second day also rescued photoreceptors. In contrast, systemic VPA treatment of Pde6b(rd10/rd10) mice (P17-P28) reduced visual function that correlated with a slight increase in photoreceptor loss. Treating Pde6b(rd10/rd10) mice earlier (P9-P21) also failed to rescue photoreceptors. Treating wild-type mice earlier (P9-P21) reduced the number of photoreceptors in VPA-treated mice by 20% compared to PBS-treated animals.

Conclusions: A single systemic dose of VPA can change retinal neurotrophic factor and rod-specific gene expression in the immature retina. Daily VPA treatment from P17 to P28 can also alter gene expression in the mature neural retina. While daily treatment with VPA could significantly reduce photoreceptor loss in the rd1 model, VPA treatment slightly accelerated photoreceptor loss in the rd10 model. The apparent rescue of photoreceptors in the rd1 model was not the result of producing more photoreceptors before degeneration. In fact, daily systemic VPA was toxic to wild-type photoreceptors when started at P9. However, the effective treatment period for Pde6b(rd1/rd1) mice (P9-P21) has significant overlap with the photoreceptor maturation period, which complicates the use of the rd1 model for testing of VPA's efficacy. In contrast, VPA treatment started after P17 did not cause photoreceptor loss in wild-type mice. Thus, the acceleration of photoreceptor loss in the rd10 model may be more relevant where both photoreceptor loss and VPA treatment (P17-P28) started when the central retina was mature.

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Effect of daily systemic valproic acid (VPA; P17–P28, late treatment) on neurotrophic factor and rod-specific gene expression in the wild-type mouse retina. A daily dose of VPA (350 mg/kg) was administered by intraperitoneal injection starting at age P17. Littermates received injections of PBS only. Gene expression, relative to PBS-injected littermates at age P28 was measured using real-time PCR for the gene of interest (Gdnf, Bdnf, Cntf, Fgf2, Mef2c, Nrl, Crx, Nr2E3, Rho, Pde6b) and was normalized to Actb expression (mean±standard deviation; n = 3/group, t test, *p<0.05, **p<0.01, relative to PBS).
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f3: Effect of daily systemic valproic acid (VPA; P17–P28, late treatment) on neurotrophic factor and rod-specific gene expression in the wild-type mouse retina. A daily dose of VPA (350 mg/kg) was administered by intraperitoneal injection starting at age P17. Littermates received injections of PBS only. Gene expression, relative to PBS-injected littermates at age P28 was measured using real-time PCR for the gene of interest (Gdnf, Bdnf, Cntf, Fgf2, Mef2c, Nrl, Crx, Nr2E3, Rho, Pde6b) and was normalized to Actb expression (mean±standard deviation; n = 3/group, t test, *p<0.05, **p<0.01, relative to PBS).

Mentions: We used an intermediate dose (350 mg/kg/day) for daily treatment of wild-type mice with VPA as it involved multiple doses delivered from age P17 to age P28. We employed SD-OCT to confirm that VPA treatment started at P17 has no effects on retinal morphology before collecting the tissue for gene expression analysis. Based on an average of 24 different OCT measurement locations from both eyes of each mouse, the average ONL thickness was exactly the same comparing VPA-treated (63±2 µm, n = 3) and PBS-treated (63±2 µm, n = 3) littermates. Examples of high-resolution B-scans of a VPA-treated and PBS-treated littermate are shown in Appendix 1. We compared the expression of several neurotrophic factors and rod photoreceptor specific genes from these same mice (Figure 3). Retinal Bdnf and Gdnf gene expression was elevated 1.6- and twofold, respectively, in the VPA-treated littermates. Retinal Cntf gene expression of VPA-treated mice was the same as their PBS-treated littermates, while Fgf2 gene expression was diminished by 25%. Most notable was a 50% reduction in endogenous Nrl gene expression by the end of the dosing period. In contrast, Crx gene expression was the same comparing VPA-treated and PBS-treated littermates. Both Rhodopsin and Pde6b gene expression in VPA-treated mice attained the same level as their PBS-treated littermates by age P28. In contrast, the NRL-dependent and rod-specific expression of the Mef2c gene was reduced by 90%. Expression of the rod-specific co-activator/co-repressor Nr2e3 gene was also reduced by 55% in VPA-treated mice compared to PBS-treated littermates (Figure 3).


Different effects of valproic acid on photoreceptor loss in Rd1 and Rd10 retinal degeneration mice.

Mitton KP, Guzman AE, Deshpande M, Byrd D, DeLooff C, Mkoyan K, Zlojutro P, Wallace A, Metcalf B, Laux K, Sotzen J, Tran T - Mol. Vis. (2014)

Effect of daily systemic valproic acid (VPA; P17–P28, late treatment) on neurotrophic factor and rod-specific gene expression in the wild-type mouse retina. A daily dose of VPA (350 mg/kg) was administered by intraperitoneal injection starting at age P17. Littermates received injections of PBS only. Gene expression, relative to PBS-injected littermates at age P28 was measured using real-time PCR for the gene of interest (Gdnf, Bdnf, Cntf, Fgf2, Mef2c, Nrl, Crx, Nr2E3, Rho, Pde6b) and was normalized to Actb expression (mean±standard deviation; n = 3/group, t test, *p<0.05, **p<0.01, relative to PBS).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4225157&req=5

f3: Effect of daily systemic valproic acid (VPA; P17–P28, late treatment) on neurotrophic factor and rod-specific gene expression in the wild-type mouse retina. A daily dose of VPA (350 mg/kg) was administered by intraperitoneal injection starting at age P17. Littermates received injections of PBS only. Gene expression, relative to PBS-injected littermates at age P28 was measured using real-time PCR for the gene of interest (Gdnf, Bdnf, Cntf, Fgf2, Mef2c, Nrl, Crx, Nr2E3, Rho, Pde6b) and was normalized to Actb expression (mean±standard deviation; n = 3/group, t test, *p<0.05, **p<0.01, relative to PBS).
Mentions: We used an intermediate dose (350 mg/kg/day) for daily treatment of wild-type mice with VPA as it involved multiple doses delivered from age P17 to age P28. We employed SD-OCT to confirm that VPA treatment started at P17 has no effects on retinal morphology before collecting the tissue for gene expression analysis. Based on an average of 24 different OCT measurement locations from both eyes of each mouse, the average ONL thickness was exactly the same comparing VPA-treated (63±2 µm, n = 3) and PBS-treated (63±2 µm, n = 3) littermates. Examples of high-resolution B-scans of a VPA-treated and PBS-treated littermate are shown in Appendix 1. We compared the expression of several neurotrophic factors and rod photoreceptor specific genes from these same mice (Figure 3). Retinal Bdnf and Gdnf gene expression was elevated 1.6- and twofold, respectively, in the VPA-treated littermates. Retinal Cntf gene expression of VPA-treated mice was the same as their PBS-treated littermates, while Fgf2 gene expression was diminished by 25%. Most notable was a 50% reduction in endogenous Nrl gene expression by the end of the dosing period. In contrast, Crx gene expression was the same comparing VPA-treated and PBS-treated littermates. Both Rhodopsin and Pde6b gene expression in VPA-treated mice attained the same level as their PBS-treated littermates by age P28. In contrast, the NRL-dependent and rod-specific expression of the Mef2c gene was reduced by 90%. Expression of the rod-specific co-activator/co-repressor Nr2e3 gene was also reduced by 55% in VPA-treated mice compared to PBS-treated littermates (Figure 3).

Bottom Line: Nrl gene expression was decreased by 50%, while Crx gene expression was not affected.Rod-specific expression of Mef2c and Nr2e3 was decreased substantially by VPA treatment, while Rhodopsin and Pde6b gene expression was normal at P28.While daily treatment with VPA could significantly reduce photoreceptor loss in the rd1 model, VPA treatment slightly accelerated photoreceptor loss in the rd10 model.

View Article: PubMed Central - PubMed

Affiliation: Control of Gene Expression Laboratory and Pediatric Retinal Research Laboratory, Eye Research Institute, Oakland University, Rochester, MI.

ABSTRACT

Purpose: The histone-deacetylase inhibitor activity of valproic acid (VPA) was discovered after VPA's adoption as an anticonvulsant. This generated speculation for VPA's potential to increase the expression of neuroprotective genes. Clinical trials for retinitis pigmentosa (RP) are currently active, testing VPA's potential to reduce photoreceptor loss; however, we lack information regarding the effects of VPA on available mammalian models of retinal degeneration, nor do we know if retinal gene expression is perturbed by VPA in a predictable way. Thus, we examined the effects of systemic VPA on neurotrophic factor and Nrl-related gene expression in the mouse retina and compared VPA's effects on the rate of photoreceptor loss in two strains of mice, Pde6b(rd1/rd1) and Pde6b(rd10/rd10) .

Methods: The expression of Bdnf, Gdnf, Cntf, and Fgf2 was measured by quantitative PCR after single and multiple doses of VPA (intraperitoneal) in wild-type and Pde6b(rd1/rd1) mice. Pde6b(rd1/rd1) mice were treated with daily doses of VPA during the period of rapid photoreceptor loss. Pde6b(rd10/rd10) mice were also treated with systemic VPA to compare in a partial loss-of-function model. Retinal morphology was assessed by virtual microscopy or spectral-domain optical coherence tomography (SD-OCT). Full-field and focal electroretinography (ERG) analysis were employed with Pde6b(rd10/rd10) mice to measure retinal function.

Results: In wild-type postnatal mice, a single VPA dose increased the expression of Bdnf and Gdnf in the neural retina after 18 h, while the expression of Cntf was reduced by 70%. Daily dosing of wild-type mice from postnatal day P17 to P28 resulted in smaller increases in Bdnf and Gdnf expression, normal Cntf expression, and reduced Fgf2 expression (25%). Nrl gene expression was decreased by 50%, while Crx gene expression was not affected. Rod-specific expression of Mef2c and Nr2e3 was decreased substantially by VPA treatment, while Rhodopsin and Pde6b gene expression was normal at P28. Daily injections with VPA (P9-P21) dramatically slowed the loss of rod photoreceptors in Pde6b(rd1/rd1) mice. At age P21, VPA-treated mice had several extra rows of rod photoreceptor nuclei compared to PBS-injected littermates. Dosing started later (P14) or dosing every second day also rescued photoreceptors. In contrast, systemic VPA treatment of Pde6b(rd10/rd10) mice (P17-P28) reduced visual function that correlated with a slight increase in photoreceptor loss. Treating Pde6b(rd10/rd10) mice earlier (P9-P21) also failed to rescue photoreceptors. Treating wild-type mice earlier (P9-P21) reduced the number of photoreceptors in VPA-treated mice by 20% compared to PBS-treated animals.

Conclusions: A single systemic dose of VPA can change retinal neurotrophic factor and rod-specific gene expression in the immature retina. Daily VPA treatment from P17 to P28 can also alter gene expression in the mature neural retina. While daily treatment with VPA could significantly reduce photoreceptor loss in the rd1 model, VPA treatment slightly accelerated photoreceptor loss in the rd10 model. The apparent rescue of photoreceptors in the rd1 model was not the result of producing more photoreceptors before degeneration. In fact, daily systemic VPA was toxic to wild-type photoreceptors when started at P9. However, the effective treatment period for Pde6b(rd1/rd1) mice (P9-P21) has significant overlap with the photoreceptor maturation period, which complicates the use of the rd1 model for testing of VPA's efficacy. In contrast, VPA treatment started after P17 did not cause photoreceptor loss in wild-type mice. Thus, the acceleration of photoreceptor loss in the rd10 model may be more relevant where both photoreceptor loss and VPA treatment (P17-P28) started when the central retina was mature.

Show MeSH
Related in: MedlinePlus