Current Stem-Cell Approaches for the Treatment of Inherited Retinal Degenerations.

Stem cells provide a promising new therapeutic approach for the treatment of multiple acquired and inherited retinal conditions. While to date, there have been numerous clinical trials examining the ability of stem cells to treat the geographic atrophy found in advanced non-neovascular age-related macular degeneration, fewer clinical trials have specifically examined stem-cell therapy for inherited retinal disease. Moreover, it remains to be seen if human stem cells will be able to regenerate the lost retinal cell populations that represent a final common pathway for most of the inherited retinal diseases, or if stem cells will secrete a neuroprotective paracrine factor that will delay progression of these diseases. Here, we will review a number of the current clinical trials, either completed or in process, that have been designed to specifically treat inherited retinal conditions. There was considerable initial concern that using human stem cells as therapeutic agents might have the potential to form benign tumors or trigger an immune response that would have deleterious effects on the patient's retina. Currently, the majority of the clinical trials reviewed share the conclusion that intraocular stem-cell approach is generally well tolerated and safe for patients. While there are some efficacy data that have been published for a few of the reviewed trials, none of the completed studies have been empowered to demonstrate statistically significant efficacy in humans.

Opposite feedbacks in the Hippo pathway for growth control and neural fate.

Signaling pathways are reused for multiple purposes in plant and animal development. The Hippo pathway in mammals and Drosophila coordinates proliferation and apoptosis via the coactivator and oncoprotein YAP/Yorkie (Yki), which is homeostatically regulated through negative feedback. In the Drosophila eye, cross-repression between the Hippo pathway kinase LATS/Warts (Wts) and growth regulator Melted generates mutually exclusive photoreceptor subtypes. Here, we show that this all-or-nothing neuronal differentiation results from Hippo pathway positive feedback: Yki both represses its negative regulator, warts, and promotes its positive regulator, melted. This postmitotic Hippo network behavior relies on a tissue-restricted transcription factor network-including a conserved Otx/Orthodenticle-Nrl/Traffic Jam feedforward module-that allows Warts-Yki-Melted to operate as a bistable switch. Altering feedback architecture provides an efficient mechanism to co-opt conserved signaling networks for diverse purposes in development and evolution.

Alterations of the CIB2 calcium- and integrin-binding protein cause Usher syndrome type 1J and nonsyndromic deafness DFNB48.

Sensorineural hearing loss is genetically heterogeneous. Here, we report that mutations in CIB2, which encodes a calcium- and integrin-binding protein, are associated with nonsyndromic deafness (DFNB48) and Usher syndrome type 1J (USH1J). One mutation in CIB2 is a prevalent cause of deafness DFNB48 in Pakistan; other CIB2 mutations contribute to deafness elsewhere in the world. In mice, CIB2 is localized to the mechanosensory stereocilia of inner ear hair cells and to retinal photoreceptor and pigmented epithelium cells. Consistent with molecular modeling predictions of calcium binding, CIB2 significantly decreased the ATP-induced calcium responses in heterologous cells, whereas mutations in deafness DFNB48 altered CIB2 effects on calcium responses. Furthermore, in zebrafish and Drosophila melanogaster, CIB2 is essential for the function and proper development of hair cells and retinal photoreceptor cells. We also show that CIB2 is a new member of the vertebrate Usher interactome.

OTX2 and CRX rescue overlapping and photoreceptor-specific functions in the Drosophila eye.

BACKGROUND: Otd-related transcription factors are evolutionarily conserved to control anterior patterning and neurogenesis. In humans, two such factors, OTX2 and CRX, are expressed in all photoreceptors from early specification through adulthood and associate with several photoreceptor-specific retinopathies. It is not well understood how these factors function independently vs. redundantly, or how specific mutations lead to different disease outcomes. It is also unclear how OTX1 and OTX2 functionally overlap during other aspects of neurogenesis and ocular development. Drosophila encodes a single Otd factor that has multiple functions during eye development. Using the Drosophila eye as a model, we tested the ability of the human OTX1, OTX2, and CRX genes, as well as several disease-associated CRX alleles, to rescue the different functions of Otd. RESULTS: Our results indicate the following: OTX2 and CRX display overlapping, yet distinct subfunctions of Otd during photoreceptor differentiation; CRX disease alleles can be functionally distinguished based on their rescue properties; and all three factors are able to rescue rhabdomeric photoreceptor morphogenesis. CONCLUSIONS: Our findings have important implications for understanding how Otx proteins have subfunctionalized during evolution, and cement Drosophila as an effective tool to unravel the molecular bases of photoreceptor pathogenesis.

Separable transcriptional regulatory domains within Otd control photoreceptor terminal differentiation events.

Orthodenticle (Otd)-related transcription factors are essential for anterior patterning and brain morphogenesis from Cnidaria to Mammals, and genetically underlie several human retinal pathologies. Despite their key developmental functions, relatively little is known regarding the molecular basis of how these factors regulate downstream effectors in a cell- or tissue-specific manner. Many invertebrate and vertebrate species encode two to three Otd proteins, whereas Drosophila encodes a single Otd protein. In the fly retina, Otd controls rhabdomere morphogenesis of all photoreceptors and regulates distinct Rhodopsin-encoding genes in a photoreceptor subtype-specific manner. Here, we performed a structure-function analysis of Otd during Drosophila eye development using in vivo rescue experiments and in vitro transcriptional regulatory assays. Our findings indicate that Otd requires at least three distinct transcriptional regulatory domains to control photoreceptor-specific rhodopsin gene expression and photoreceptor morphogenesis. Our results also uncover a previously unknown role for Otd in preventing co-expression of sensory receptors in blue vs. green-sensitive R8 photoreceptors. Sequence analysis indicates that many of the transcriptional regulatory domains identified here are conserved in multiple Diptera Otd-related proteins. Thus, these studies provide a basis for identifying shared molecular pathways involved in a wide range of developmental processes.

Monte Carlo studies of isomers, structures, and properties in benzene-cyclohexane clusters: computation strategy and application to the dimer and trimer, (C6H6)(C6H12)n, n = 1-2.

Low-temperature isomeric energies, structures, and properties of benzene-cyclohexane clusters are investigated via Monte Carlo simulations. The Monte Carlo strategy is first documented and then applied to (C(6)H(6))(C(6)H(12)) and (C(6)H(6))(C(6)H(12))(2) using four different potential energy surfaces. Results identify a single parallel-displaced dimer isomer. MP2 optimizations and frequency calculations support the Monte Carlo dimer structure and identify the van der Waals mode observed in vibronic spectra. Caloric simulations identify two temperatures where structural transitions occur and imply an experimental temperature below 10 K for dimers in cold supersonic expansions. The (C(6)H(6))(C(6)H(12))(2) studies identify eight independent trimer isomers: three form parallel-stacked (sandwich) arrangements with the two cyclohexane moieties related through a D(6)(h) transformation. The remaining five trimer isomers are trigonal, with no overall symmetry. Caloric studies indicate that the sandwich and trigonal isomeric classes coexist independently below 60 K, consistent with trimer vibronic spectra that contain two independent van der Waals progressions.