Debbie S. Kuo

COL4A1 and COL4A2 Mutations and Disease: Insights into Pathogenic Mechanisms and Potential Therapeutic Targets

Heterotrimers composed of collagen type IV alpha 1 (COL4A1) and alpha 2 (COL4A2) constitute one of the most abundant components of nearly all basement membranes. Accordingly, mutations in COL4A1 or COL4A2 are pleiotropic and contribute to a broad spectrum of disorders, including myopathy, glaucoma and hemorrhagic stroke. Here, we summarize the contributions of COL4A1 and COL4A2 mutations in human disease, integrate knowledge gained from model organisms and evaluate the implications for pathogenic mechanisms and therapeutic approaches.

COL4A2 mutation associated with familial porencephaly and small-vessel disease

Familial porencephaly, leukoencephalopathy and small-vessel disease belong to the spectrum of disorders ascribed to dominant mutations in the gene encoding for type IV collagen alpha-1 (COL4A1). Mice harbouring mutations in either Col4a1 or Col4a2 suffer from porencephaly, hydrocephalus, cerebral and ocular bleeding and developmental defects. We observed porencephaly and white matter lesions in members from two families that lack COL4A1 mutations. We hypothesized that COL4A2 mutations confer genetic predisposition to porencephaly, therefore we sequenced COL4A2 in the family members and characterized clinical, neuroradiological and biochemical phenotypes. Genomic sequencing of COL4A2 identified the heterozygous missense G1389R in exon 44 in one family and the c.3206delC change in exon 34 leading to frame shift and premature stop, in the second family. Fragmentation and duplication of epidermal basement membranes were observed by electron microscopy in a c.3206delC patient skin biopsy, consistent with abnormal collagen IV network. Collagen chain accumulation and endoplasmic reticulum (ER) stress have been proposed as cellular mechanism in COL4A1 mutations. In COL4A2 3206delC fibroblasts we detected increased rates of apoptosis and no signs of ER stress. Mutation phenotypes varied, including porencephaly, white matter lesions, cerebellar and optic nerve hypoplasia and unruptured carotid aneurysm. In the second family however, we found evidence for additional factors contributing to the phenotype. We conclude that dominant COL4A2 mutations are a novel major risk factor for familial cerebrovascular disease, including porencephaly and small-vessel disease with reduced penetrance and variable phenotype, which might also be modified by other contributing factors.

The short-term effects of exercise on intraocular pressure, choroidal thickness and axial length.

Purpose To explore ocular changes in healthy people after exercise. Methods Twenty five volunteers underwent exercise for 15 minutes on a treadmill. Measurements of choroidal thickness, intraocular pressure (IOP), ocular biometry, and blood pressure were taken before and after exercise. Enhanced Depth Imaging optical coherence tomography (EDI-OCT) was used to measure choroidal thickness at the fovea. Intraocular pressure (IOP) was measured by Goldmann applanation tonometry. Ocular biometric measures were collected using A scan ultrasound. Blood pressure was measured concurrently with the acquisition of the scans. Results Twenty five volunteers (25 eyes) with a mean age of 25.44±3.25 years were measured. There was a significant increase in systolic and diastolic pressure after exercise (P<0.05). The IOP showed a significant decrease after exercise (P<0.05). However there was no significant difference in the mean choroidal thickness, ocular axial length, anterior chamber depth, lens thickness, or vitreous length before and after exercise measurements (P>0.05). Conclusion There was a significant decrease in IOP from exercise without a change in choroidal thickness and ocular biometric measures. IOP and choroidal thickness were not correlated, suggesting that the IOP decrease from exercise is not due to changes in choridal thickness.

Controlling the number of melanopsin-containing retinal ganglion cells by early light exposure.

A small percentage of retinal ganglion cells (RGCs) express melanopsin and are intrinsically photosensitive (ipRGCs). Whether light can affect the development of ipRGCs is not clear. In the rat retina, we found constant light exposure during the first postnatal week significantly increased the number of melanopsin immunopositive ipRGCs. This increase was durable and specific for melanopsin immunopositive ipRGCs. BrdU labeling showed no proliferation of the melanopsin immunopositive ipRGCs during constant light exposure. Retrograde labeling from the superior colliculus showed that no other types of RGCs were induced to express melanopsin. Light exposure was effective in increasing melanopsin immunopositive ipRGCs only when it coincided with the apoptotic phase of RGC development. However, daily intravitreous injection of tetrodotoxin, blocking action potentials, abolished the light induced increase of melanopsin immunopositive ipRGCs. These findings indicate that early light exposure can increase the number of melanopsin immunopositive ipRGCs through a process dependent on intrinsic photosensitive spiking activity. Furthermore, the increase of melanopsin immunopositive ipRGCs is potentially induced by apoptosis suppression in ipRGCs or enhanced expression of melanopsin.

Characterization of a variant of gap junction protein α8 identified in a family with hereditary cataract

Purpose Congenital cataracts occur in isolation in about 70% of cases or are associated with other abnormalities such as anterior segment dysgenesis and microphthalmia. We identified a three-generation family in the University of California San Francisco glaucoma clinic comprising three individuals with congenital cataracts and aphakic glaucoma, one of whom also had microphthalmia. The purpose of this study was to identify a possible causative mutation in this family and to investigate its pathogenesis. Methods We performed exome sequencing and identified a putative mutation in gap junction protein α8 (GJA8). We used PCR and DNA sequencing of GJA8 in affected and unaffected members of the pedigree to test segregation of the variant with the phenotype. We tested cellular distribution and function of the variant protein by immunofluorescence and intercellular transfer of Neurobiotin in transiently transfected HeLa cells. Results Exome sequencing revealed a variant in GJA8 (c.658A>G) encoding connexin50 (Cx50) that resulted in a missense change (p.N220D) in transmembrane domain 4. The variant was present in all three affected family members, but was also present in the probands grandfather who was reported to be unaffected. The mutant protein localized to the plasma membrane and supported intercellular Neurobiotin transfer in HeLa cells. Conclusions We identified a variant in transmembrane domain 4 of Cx50 in a family with autosomal dominant congenital cataracts. This variant has been previously identified in other cataract cohorts, but it is also present in unaffected individuals. Our study demonstrates that the mutant protein localized to the plasma membrane and formed functional intercellular channels. These data suggest that GJA8 c.658A>G is most likely a benign rare variant.

Congenital solitary eyelid trichoepithelioma.

Trichoepitheliomas are unusual, benign tumors of hair follicle origin. They may present in children and adults as acquired lesions on the body and face, but they rarely involve the eyelids. Solitary trichoepitheliomas have not previously been reported in infants. We present a first report of congenital solitary eyelid trichoepithelioma in an infant and review the classification of trichoepitheliomas.