Case Report: Retinal Toxicity Secondary to Ritonavir.

SIGNIFICANCE: Because patients with HIV have increased life expectancies with the advent of new drug therapies, complications from iatrogenic syndromes such as drug toxicity can occur. Ritonavir-induced retinal toxicity is one such complication but has rarely been reported in the literature. PURPOSE: This case report describes a patient with bilateral maculopathy and bone spicule-like pigmentary changes in the midperipheral retina due to ritonavir use. In addition, novel optical coherence tomography findings are described. CASE REPORT: A 53-year-old man presented with gradual-onset blurry vision and difficulty seeing at night. He had been diagnosed as having HIV infection 19 years prior and had previously taken ritonavir for 7 years as part of highly active antiretroviral therapy. Best-corrected acuities were 20/30 in the right eye and 20/25 in the left eye. Clinical examination revealed a subtle annular pattern of retinal pigment epithelium mottling around the fovea and bone spicule-like pigment changes in the midperiphery of both retinas. Optical coherence tomography imaging revealed abnormal subfoveal hyperreflectivity of the ellipsoid zone with relative attenuation centrally, annular parafoveal ellipsoid zone loss, and punctate hyperreflectivity within the ellipsoid zone more eccentrically. Fundus autofluorescence in both eyes showed annular hyperautofluorescence in the parafoveal region, geographic hyperautofluorescence in the areas underlying the midperipheral pigmentary changes, and discrete patches of hyperautofluorescence along the far inferotemporal arcades in areas that appeared normal with biomicroscopy. A diagnosis of retinal toxicity secondary to ritonavir use was made based on the patient's history and clinical examination. CONCLUSIONS: Ritonavir retinal toxicity seems to be an uncommon adverse event that can cause decreased visual function. This case report provides further evidence of the retinal toxicity and reviews the reported clinical and optical coherence tomography manifestations of the disease.

Constitutive protein kinase A activity in osteocytes and late osteoblasts produces an anabolic effect on bone.

Osteocytes have been implicated in the control of bone formation. However, the signal transduction pathways that regulate the biological function of osteocytes are poorly defined. Limited evidence suggests an important role for the Gs/cAMP pathway in osteocyte function. In the present study, we explored the hypothesis that cAMP-dependent kinase A (PKA) activation in osteocytes plays a key role in controlling skeletal homeostasis. To test this hypothesis, we mated mice harboring a Cre-conditional, mutated PKA catalytic subunit allele that encodes a constitutively active form of PKA (CαR) with mice expressing Cre under the control of the osteocyte-specific promoter, DMP1. This allowed us to direct the expression of CαR to osteocytes in double transgenic progeny. Examination of Cre expression indicated that CαR was also expressed in late osteoblasts. Cortical and trabecular bone parameters from 12-week old mice were determined by µCT. Expression of CαR in osteocytes and late osteoblasts altered the shape of cortical bone proximal to the tibia-fibular junction (TFJ) and produced a significant increase in its size. In trabecular bone of the distal femur, fractional bone volume, trabecular number, and trabecular thickness were increased. These increases were partially the results of increased bone formation rates (BFRs) on the endosteal surface of the cortical bone proximal to the TFJ as well as increased BFR on the trabecular bone surface of the distal femur. Mice expressing CαR displayed a marked increase in the expression of osteoblast markers such as osterix, runx2, collagen 1α1, and alkaline phosphatase (ALP). Interestingly, expression of osteocyte marker gene, DMP1, was significantly up-regulated but the osteocyte number per bone area was not altered. Expression of SOST, a presumed target for PKA signaling in osteocytes, was significantly down-regulated in females. Importantly, no changes in bone resorption were detected. In summary, constitutive PKA signaling in osteocytes and late osteoblasts led to a small expansion of the size of the cortical bone proximal to the TFJ and an increase in trabecular bone in female mice. This was associated with down-regulation of SOST and up-regulation of several osteoblast marker genes. Activation of the PKA pathway in osteocytes and late osteoblasts is sufficient for the initiation of an anabolic skeletal response.

Cyclic AMP signaling in bone marrow stromal cells has reciprocal effects on the ability of mesenchymal stem cells to differentiate into mature osteoblasts versus mature adipocytes.

Stimulatory G protein-mediated cAMP signaling is intimately involved in skeletal homeostasis. However, limited information is available on the role of the cAMP signaling in regulating the differentiation of mesenchymal stem cells into mature osteoblasts and adipocytes. To investigate this, we treated primary mouse bone marrow stromal cells (BMSCs) with forskolin to stimulate cAMP signaling and determined the effect on osteoblast and adipocyte differentiation. Exposure of differentiating osteoblasts to forskolin markedly inhibited progression to the late stages of osteoblast differentiation, and this effect was replicated by continuous exposure to PTH. Strikingly, forskolin activation of cAMP signaling in BMSCs conditioned mesenchymal stem cells (MSCs) to undergo increased osteogenic differentiation and decreased adipogenic differentiation. PTH treatment of BMSCs also enhanced subsequent osteogenesis, but promoted an increased adipogenesis as well. Thus, activation of cAMP signaling alters the lineage commitment of MSCs, favoring osteogenesis at the expense of adipogenesis.

Blockade of receptor-activated G(i) signaling in osteoblasts in vivo leads to site-specific increases in cortical and cancellous bone formation.

Osteoblasts play a critical role in the maintenance of bone mass through bone formation and regulation of bone resorption. Targeted expression of a constitutively active engineered G(i)-coupled G protein-coupled receptor (GPCR) to osteoblasts in vivo leads to severe osteopenia. However, little is known about the role of endogenous receptor-mediated G(i) signaling in regulating osteoblast function. In this study, we investigated the skeletal effects of blocking G(i)-coupled signaling in osteoblasts in vivo. This was accomplished by transgenic expression of the catalytic subunit of pertussis toxin (PTX) under control of the collagen Iα 2.3-kb promoter. These mice, designated Col1(2.3)(+)/PTX(+), showed increased cortical thickness at the femoral midshaft at 12 weeks of age. This correlated with increased periosteal bone formation associated with expanded mineralizing surface observed in 8-week-old mice of both genders. The cancellous bone phenotype of the Col1(2.3)(+)/PTX(+) mice was sexually dimorphic, with increases in fractional bone volume at the distal femur seen only in females. Similarly, while cancellous bone-formation rates were unchanged in males, they could not be quantified for female Col1(2.3)(+)/PTX(+) mice owing to the disorganized nature of the labeling pattern, which was consistent with rapid formation of woven bone. Alterations in osteoclast activity did not appear to participate in the phenotype. These data demonstrate that G(i)-coupled signaling by GPCRs endogenous to osteoblasts plays a complex role in the regulation of bone formation in a manner that is dependent on both gender and the anatomic site within bone.

Ligand-mediated activation of an engineered gs g protein-coupled receptor in osteoblasts increases trabecular bone formation.

Age-dependent changes in skeletal growth play important roles in regulating skeletal expansion and in the course of many diseases affecting bone. How G protein-coupled receptor (GPCR) signaling affects these changes is poorly understood. Previously, we described a mouse model expressing Rs1, an engineered receptor with constitutive G(s) activity. Rs1 expression in osteoblasts from gestation induced a dramatic age-dependent increase in trabecular bone with features resembling fibrous dysplasia; however, these changes were greatly minimized if Rs1 expression was delayed until after puberty. To further investigate whether ligand-induced activation of the G(s)-GPCR pathway affects bone formation in adult mice, we activated Rs1 in adult mice with the synthetic ligand RS67333 delivered continuously via an osmotic pump or intermittently by daily injections. We found that osteoblasts from adult animals can be stimulated to form large amounts of bone, indicating that adult mice are sensitive to the dramatic bone- forming actions of G(s) signaling in osteoblasts. In addition, our results show that intermittent and continuous activation of Rs1 led to structurally similar but quantitatively different degrees of trabecular bone formation. These results indicate that activation of a G(s)-coupled receptor in osteoblasts of adult animals by either intermittent or continuous ligand administration can increase trabecular bone formation. In addition, osteoblasts located at the bone epiphyses may be more responsive to G(s) signaling than osteoblasts at the bone diaphysis. This model provides a powerful tool for investigating the effects of ligand-activated G(s)-GPCR signaling on dynamic bone growth and remodeling.