Sandra Chang

Oligodendrocyte-Encoded HIF Function Couples Postnatal Myelination and White Matter Angiogenesis

Myelin sheaths provide critical functional and trophic support for axons in white matter tracts of the brain. Oligodendrocyte precursor cells (OPCs) have extraordinary metabolic requirements during development as they differentiate to produce multiple myelin segments, implying that they must first secure adequate access to blood supply. However, mechanisms that coordinate myelination and angiogenesis are unclear. Here, we show that oxygen tension, mediated by OPC-encoded hypoxia-inducible factor (HIF) function, is an essential regulator of postnatal myelination. Constitutive HIF1/2α stabilization resulted in OPC maturation arrest through autocrine activation of canonical Wnt7a/7b. Surprisingly, such OPCs also show paracrine activity that induces excessive postnatal white matter angiogenesis in vivo and directly stimulates endothelial cell proliferation in vitro. Conversely, OPC-specific HIF1/2α loss of function leads to insufficient angiogenesis in corpus callosum and catastrophic axon loss. These findings indicate that OPC-intrinsic HIF signaling couples postnatal white matter angiogenesis, axon integrity, and the onset of myelination in mammalian forebrain.

Astrocyte-encoded positional cues maintain sensorimotor circuit integrity

Astrocytes, the most abundant cells in the central nervous system, promote synapse formation and help to refine neural connectivity. Although they are allocated to spatially distinct regional domains during development, it is unknown whether region-restricted astrocytes are functionally heterogeneous. Here we show that postnatal spinal cord astrocytes express several region-specific genes, and that ventral astrocyte-encoded semaphorin 3a (Sema3a) is required for proper motor neuron and sensory neuron circuit organization. Loss of astrocyte-encoded Sema3a leads to dysregulated α-motor neuron axon initial segment orientation, markedly abnormal synaptic inputs, and selective death of α- but not of adjacent γ-motor neurons. In addition, a subset of TrkA+ sensory afferents projects to ectopic ventral positions. These findings demonstrate that stable maintenance of a positional cue by developing astrocytes influences multiple aspects of sensorimotor circuit formation. More generally, they suggest that regional astrocyte heterogeneity may help to coordinate postnatal neural circuit refinement.

Topical treatment with thiazolidinediones, activators of peroxisome proliferator-activated receptor-γ, normalizes epidermal homeostasis in a murine hyperproliferative disease model

Abstract:  In a murine model of epidermal hyperplasia reproducing some of the abnormalities of several common skin disorders, we previously demonstrated the antiproliferative and pro‐differentiating effects of peroxisome proliferator‐activated receptor (PPAR)α, PPARβ/δ, and liver X receptor activators. Unlike other subgroups of PPAR activators, thiazolidinediones (TZDs), a family of PPARγ ligands, did not inhibit keratinocyte proliferation in normal murine skin. Here, we studied the effects of two TZDs, namely ciglitazone (10 mM) and troglitazone (1 mM), in the same murine model where epidermal hyperproliferation was reproduced by repeated barrier abrogation with tape stripping. Topical treatment with ciglitazone and troglitazone resulted in a marked and significant decrease in epidermal thickness. Furthermore, in all TZD‐treated groups, we observed a significant decrease in keratinocyte proliferation using proliferating cell nuclear antigen, 5‐bromo‐2′‐deoxyuridine, and tritiated thymidine incorporation. However, using the terminal deoxynucleotidyl transferase‐mediated dUTP nick end‐labeling assay, we found no difference in apoptosis between different treatments, emphasizing that it is the antiproliferative role of these activators that accounts for the decrease of epidermal thickness. Finally, using immunohistochemical methods, we determined the effects of ciglitazone on keratinocyte differentiation in this hyperproliferative model. We observed an increased expression of involucrin and filaggrin following ciglitazone treatment, suggesting a pro‐differentiating action of TZDs in this model. In summary, topical TZDs significantly reduce epidermal keratinocyte proliferation while promoting differentiation in a murine model of hyperproliferative epidermis. Together, these results suggest that in addition to their metabolic effects currently in use in the treatment of type 2 diabetes, topical TZDs could be considered as potential alternative therapeutic agents in hyperproliferative skin diseases such as psoriasis.

Development and progression of alopecia in the vitamin D receptor null mouse

Humans with selected mutations in the vitamin D receptor (VDR) and mouse models lacking VDR develop alopecia. Mice null for the Vdr gene are born with a normal coat of hair, but fail to initiate normal hair follicle cycling. In this study, we examined the morphology of the hair follicle of the Vdr null mouse during days 13–22 when the hair follicle normally initiates and completes the first catagen. We then explored the possibility that the abnormality in hair follicle cycling was associated with abnormal expression of hairless (Hr), a putative transcriptional regulator known to regulate hair follicle cycling and recently shown to regulate VDR transcriptional activity. Our results demonstrate the progressive deterioration of the hair follicle through catagen. Comparable to VDR, Hr was found in the basal cells of the epidermis and ORS of the hair follicle. However, Hr was also found in the IRS and matrix of the follicle, regions with little or no VDR. Hr levels increased during catagen, reaching a peak by day 19. Levels of Hr were greater in the Vdr null mice compared to wildtype controls, results confirmed by quantitative RT‐PCR. We conclude that lack of VDR causes disruption of hair follicle structure during the first catagen resulting in failure of subsequent hair follicle cycling. These changes are associated with increased expression of Hr, suggesting a role for VDR in regulating Hr expression. Both Hr and VDR are required for normal hair follicle cycling. J. Cell. Physiol. 207: 340–353, 2006.

Phosphatidylinositol-4-phosphate 5-kinase 1α Mediates Extracellular Calcium-induced Keratinocyte Differentiation

Extracellular calcium (Cao) is a major regulator of keratinocyte differentiation, but the mechanism is unclear. Phosphatidylinositol-4-phosphate 5-kinase 1alpha (PIP5K1alpha) is critical in synthesizing phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2]. In this study, we sought to determine whether PIP5K1alpha plays a role in mediating the ability of Cao to induce keratinocyte differentiation. We found that treatment of human keratinocytes in culture with Cao resulted in increased PIP5K1alpha level and activity, as well as PI(4,5)P2 level, binding of phosphatidylinositol 3,4,5-triphosphate [PI(3,4,5)P3] to and activation of phospholipase C-gamma1 (PLC-gamma1), with the resultant increase in inositol 1,4,5-trisphosphate (IP3) and intracellular calcium (Cai). Knockdown of PIP5K1alpha in human keratinocytes blocked Cao-induced increases in the binding of PI(3,4,5)P3 to PLC-gamma1; PLC-gamma1 activity; levels of PI(4,5)P2, IP3, and Cai; and induction of keratinocyte differentiation markers. Coimmunoprecipitation and confocal studies revealed that Cao stimulated PIP5K1alpha recruitment to the E-cadherin-catenin complex in the plasma membrane. Knockdown of E-cadherin or beta-catenin blocked Cao-induced activation of PIP5K1alpha. These results indicate that after Cao stimulation PIP5K1alpha is recruited by the E-cadherin-catenin complex to the plasma membrane where it provides the substrate PI(4,5)P2 for both PI3K and PLC-gamma1. This signaling pathway is critical for Cao-induced generation of the second messengers IP3 and Cai and keratinocyte differentiation.

Chinese herbal medicine (Tuhuai extract) exhibits topical anti-proliferative and anti-inflammatory activity in murine disease models

Abstract:  While psoriasis is one of the most common skin disorders in humans, effective, safe and inexpensive treatments are still largely unavailable. Chinese herbal medicine (CHM) has been used for centuries for treating psoriasis and several reports claim that systemic administration of one such CHM, Tuhuai, mainly composed of flos sophorae, smilax glabra roxb and licorice, is effective in psoriasis. However, the mechanisms by which this CHM improves psoriasis are not yet clear. Two universal features of psoriasis are epidermal hyperplasia and inflammation. Moreover, drugs that specifically inhibit epidermal hyperplasia and/or inflammation are widely used to treat psoriasis. Here, we investigated whether topical applications of Tuhuai extract exhibit anti‐proliferative and anti‐inflammatory activities in two murine models of inflammatory dermatoses. To assess Tuhuai’s potential anti‐proliferative effect, we disrupted epidermal barrier function twice‐daily for 4 days in normal hairless mice followed by topical applications of either 1% Tuhuai extract or Vehicle to both flanks immediately after each barrier perturbation. Changes in epidermal proliferation and apoptosis were evaluated by immunohistochemistry and TUNEL staining. To assess the anti‐inflammatory effects of Tuhuai, both irritant (phorbol ester) and acute allergic contact dermatitis (oxazolone) models were used. Whereas topical Tuhuai extract did not alter epidermal proliferation or induce irritation in normal skin, it both reduced epidermal hyperplasia in the epidermal hyperproliferative model, and reduced inflammation in both irritant and allergic contact dermatitis models. As topical Tuhuai extract exhibits anti‐proliferative and anti‐inflammatory properties in a variety of human models of inflammatory dermatoses, Tuhuai could provide an effective, relatively safe and inexpensive therapeutic alternative for the treatment of inflammatory dermatoses, including psoriasis.

Dysregulation of locus coeruleus development in congenital central hypoventilation syndrome

Human congenital central hypoventilation syndrome (CCHS), resulting from mutations in transcription factor PHOX2B, manifests with impaired responses to hypoxemia and hypercapnia especially during sleep. To identify brainstem structures developmentally affected in CCHS, we analyzed two postmortem neonatal-lethal cases with confirmed polyalanine repeat expansion (PARM) or Non-PARM (PHOX2B∆8) mutation of PHOX2B. Both human cases showed neuronal losses within the locus coeruleus (LC), which is important for central noradrenergic signaling. Using a conditionally active transgenic mouse model of the PHOX2B∆8 mutation, we found that early embryonic expression (<E10.5) caused failure of LC neuronal specification and perinatal respiratory lethality. In contrast, later onset (E11.5) of PHOX2B∆8 expression was not deleterious to LC development and perinatal respiratory lethality was rescued, despite failure of chemosensor retrotrapezoid nucleus formation. Our findings indicate that early-onset mutant PHOX2B expression inhibits LC neuronal development in CCHS. They further suggest that such mutations result in dysregulation of central noradrenergic signaling, and therefore, potential for early pharmacologic intervention in humans with CCHS.

Hypercalcemia and Overexpression of CYP27B1 in a Patient With Nephrogenic Systemic Fibrosis: Clinical Vignette and Literature Review†‡

Nephrogenic systemic fibrosis (NSF) is a disease of thickened, hard, hyperpigmented skin lesions with or without systemic fibrosis occurring in patients with renal insufficiency and associated with the administration of gadolinium‐containing contrast. The pathogenesis of this disease is unclear, and there is no definitive treatment. We describe a 71‐yr‐old patient with stable chronic lymphocytic leukemia (CLL), end‐stage renal disease (ESRD), and NSF who presented with hypercalcemia in 2006. Before onset of renal insufficiency in 2002, serum calcium, phosphorus, and PTH levels were normal. In 2004, the patient began hemodialysis, and he was diagnosed with NSF in 2005, shortly after undergoing an MRI with gadolinium contrast administration. Over the next 6 mo, albumin‐corrected serum total calcium levels rose from 9.9 to 13.1 mg/dl (normal range, 8.5–10.5 mg/dl) with normal serum phosphorus levels. On admission in September 2006, 1,25‐dihydroxyvitamin D [1,25(OH)2D] levels were elevated at 130.7 pg/ml (normal range, 25.1–66.1 pg/ml). Biopsy of an NSF lesion showed increased 25‐hydroxyvitamin D3–1‐α hydroxylase (CYP27B1) immunostaining compared with the biopsy from a normal control. This is the first reported association of NSF with hypercalcemia caused by elevated 1,25(OH)2D levels. This metabolic disturbance should be sought in future cases to determine a connection between NSF, 1,25(OH)2D metabolism, and CYP27B1 activation in the skin, which may shed light on the pathogenesis of this unusual local and systemic fibrosing disorder.

Single-cell in situ transcriptomic map of astrocyte cortical layer diversity

During organogenesis, patterns and gradients of gene expression underlie organization and diversified cell specification to generate complex tissue architecture. While the cerebral cortex is organized into six excitatory neuronal layers, it is unclear whether glial cells are diversified to mimic neuronal laminae or show distinct layering. To determine the molecular architecture of the mammalian cortex, we developed a high content pipeline that can quantify single-cell gene expression in situ. The Large-area Spatial Transcriptomic (LaST) map confirmed expected cortical neuron layer organization and also revealed a novel neuronal identity signature. Screening 46 candidate genes for astrocyte diversity across the cortex, we identified grey matter superficial, mid and deep astrocyte identities in gradient layer patterns that were distinct from neurons. Astrocyte layers formed in early postnatal cortex and mostly persisted in adult mouse and human cortex. Mutations that shifted neuronal post-mitotic identity or organization were sufficient to alter glial layering, indicating an instructive role for neuronal cues. In normal mouse cortex, astrocyte layer patterns showed area diversity between functionally distinct cortical regions. These findings indicate that excitatory neurons and astrocytes cells are organized into distinct lineage-associated laminae, which give rise to higher order neuroglial complexity of cortical architecture.