Epidermal Lipid Research Articles

Impaired Epidermal Ceramide Synthesis Causes Autosomal Recessive Congenital Ichthyosis and Reveals the Importance of Ceramide Acyl Chain Length

The barrier function of the human epidermis is supposed to be governed by lipid composition and organization in the stratum corneum. Disorders of keratinization, namely ichthyoses, are typically associated with disturbed barrier activity. Using autozygosity mapping and exome sequencing, we have identified a homozygous missense mutation in CERS3 in patients with congenital ichthyosis characterized by collodion membranes at birth, generalized scaling of the skin, and mild erythroderma. We demonstrate that the mutation inactivates ceramide synthase 3 (CerS3), which is synthesized in skin and testis, in an assay of N-acylation with C26-CoA, both in patient keratinocytes and using recombinant mutant proteins. Moreover, we show a specific loss of ceramides with very long acyl chains from C26 up to C34 in terminally differentiating patient keratinocytes, which is in line with findings from a recent CerS3-deficient mouse model. Analysis of reconstructed patient skin reveals disturbance of epidermal differentiation with an earlier maturation and an impairment of epidermal barrier function. Our findings demonstrate that synthesis of very long chain ceramides by CerS3 is a crucial early step for the skin barrier formation and link disorders presenting with congenital ichthyosis to defects in sphingolipid metabolism and the epidermal lipid architecture.

Activation of TLR3 in Keratinocytes Increases Expression of Genes Involved in Formation of the Epidermis, Lipid Accumulation, and Epidermal Organelles

Injury to the skin, and the subsequent release of non-coding double-stranded RNA from necrotic keratinocytes, has been identified as an endogenous activator of Toll-like receptor 3 (TLR3). Since changes in keratinocyte growth and differentiation follow injury, we hypothesized that TLR3 might trigger some elements of the barrier repair program in keratinocytes. Double-stranded RNA was observed to induce TLR3-dependent increases in human keratinocyte mRNA abundance for ABCA12 (ATP-binding cassette, sub-family A, member 12), glucocerebrosidase, acid sphingomyelinase, and transglutaminase 1. Additionally, treatment with double-stranded RNA resulted in increases in sphingomyelin and morphologic changes including increased epidermal lipid staining by oil-red O and TLR3-dependent increases in lamellar bodies and keratohyalin granules. These observations show that double-stranded RNA can stimulate some events in keratinocytes that are important for skin barrier repair and maintenance.

Effect of ethnicity, gender and age on the amount and composition of residual skin surface components derived from sebum, sweat and epidermal lipids

Background/purposeThe superficial layer on the skin surface, known as the acid mantle, comprises a mixture of sebum, sweat, corneocyte debris and constituents of natural moisturizing factor. Thus, the phrase ‘residual skin surface components’ (RSSC) is an appropriate term for the mixture of substances recovered from the skin surface. There is no general agreement about the effects of ethnicity, gender and age on RSSC. The aim of this human volunteer study was to evaluate RSSC in relation to ethnicity, gender and age. A suitable acquisition medium for RSSC collection was identified and samples of RSSC were subsequently analysed using gas chromatography-mass spectrometry (GC-MS) and gravimetry.MethodsA total of 315 volunteers participated in the study from a range of self-declared ethnic backgrounds. Six acquisition media were compared to determine the most suitable media for RSSC collection. The effect of age, gender and ethnicity on RSSC collection was evaluated by gravimetric analysis while GC-MS was used to determine the composition of RSSC.ResultsOf the six candidate materials assessed, cigarette paper provided the most practical and reproducible sample acquisition medium. There was no significant difference in the amount of RSSC collected when based on gender and ethnicity and no significant correlation between RSSC recovery and age. Up to 49 compounds were detected from human RSSC when analysed by GC-MS.ConclusionsThe results of the present study suggest that RSSC can be effectively collected using cigarette paper and analysed by GC-MS. Ethnicity, gender and age had no significant impact on the quantity of RSSC recovered from the skin surface.

A pilot study demonstrating a non‐invasive method for the measurement of protein turnover in skin disorders: application to psoriasis

BackgroundPrevious studies of epidermal kinetics in psoriasis have relied on invasive biopsy procedures or the use of radioactive labels. We previously developed a non-invasive method for measuring keratin synthesis in human skin using deuterated water labeling, serial collection of tape strips and measurement of deuterium enrichment in protein by mass spectrometry. This powerful method can be applied to measure other skin proteins and lipids collected by tape stripping. Here, for the first time, we apply this technique to investigate the epidermal kinetics of psoriasis, the first step in defining a kinetic profile for normal skin versus activated or quiescent psoriatic skin.MethodsPsoriatic subjects were given 2H2O orally as twice-daily doses for 16–38 days. Affected and unaffected skin was sampled by tape stripping and washing (modified Pachtman method). Proteins were isolated from the tape strips by a method that enriches for keratin. Turnover times were determined by gas chromatography/mass spectrometry. Kinetic data were compared to transepidermal water loss (TEWL).ResultsDeuterium-labeled protein from lesional psoriatic skin appeared at the skin surface within 3–8 days of label administration, whereas labeled protein from non-lesional skin requires 10–20 days to appear. Psoriatic skin had similar rate of growth despite varying anatomic location. Proteins recovered from tape strips were identified by nanoscale liquid chromatography/tandem mass spectrometry. Isolated peptides were >98% from keratin in uninvolved skin and >72% keratin in psoriatic skin. Revealing that one-quarter of all newly synthesized proteins in psoriatic skin are antimicrobial defense and other immune-related proteins. TEWL values were greater in lesional than non-lesional skin, suggesting barrier compromise in psoriatic skin despite increased clinical thickness.ConclusionsThis simple, elegant, and non-invasive method for measuring epidermal protein synthesis, which can also be adapted to measure epidermal lipids, provides a metric that may reveal new insights into the mechanisms and dynamic processes underlying psoriasis and may also provide an objective scale for determining response to therapeutic agents in pre-clinical and clinical trials. This opens a pathway to the non-invasive study of kinetics of protein formation in psoriasis or other skin diseases.

Mutations in CERS3 Cause Autosomal Recessive Congenital Ichthyosis in Humans

Autosomal recessive congenital ichthyosis (ARCI) is a rare genetic disorder of the skin characterized by abnormal desquamation over the whole body. In this study we report four patients from three consanguineous Tunisian families with skin, eye, heart, and skeletal anomalies, who harbor a homozygous contiguous gene deletion syndrome on chromosome 15q26.3. Genome-wide SNP-genotyping revealed a homozygous region in all affected individuals, including the same microdeletion that partially affects two coding genes (ADAMTS17, CERS3) and abolishes a sequence for a long non-coding RNA (FLJ42289). Whereas mutations in ADAMTS17 have recently been identified in autosomal recessive Weill-Marchesani-like syndrome in humans and dogs presenting with ophthalmologic, cardiac, and skeletal abnormalities, no disease associations have been described for CERS3 (ceramide synthase 3) and FLJ42289 so far. However, analysis of additional patients with non-syndromic ARCI revealed a splice site mutation in CERS3 indicating that a defect in ceramide synthesis is causative for the present skin phenotype of our patients. Functional analysis of patient skin and in vitro differentiated keratinocytes demonstrated that mutations in CERS3 lead to a disturbed sphingolipid profile with reduced levels of epidermis-specific very long-chain ceramides that interferes with epidermal differentiation. Taken together, these data present a novel pathway involved in ARCI development and, moreover, provide the first evidence that CERS3 plays an essential role in human sphingolipid metabolism for the maintenance of epidermal lipid homeostasis.

Comparison of three different sampling methods for canine skin lipids

Epidermal lipids are of major interest in dermatological research, especially in canine atopic dermatitis. Owing to the existence of several sampling methods, the interpretation of study results is often complicated. This study aimed to compare three different sampling methods and to establish a minimally invasive method for collecting canine epidermal lipids. Skin samples from five dogs with no obvious skin abnormalities were taken from the caudal back and the inguinal region postmortem. Samples consisted of heat-separated epidermis of three skin biopsies, three scrapes and three skin scrubs. Lipids were analysed by high-performance thin-layer chromatography; the resulting bands were identified by using corresponding standards, retardation factors and mass spectrometry. The influences of the sampling method, the body site and the ceramide standards were investigated. Between body sites, significant differences were found for cholesterol sulphate, cholesteryl esters and triglycerides. Significant differences between sampling methods were detected for all lipid fractions except for cholesterol sulphate and glucosylceramides within the lipid profile, and for at least four ceramide classes within the ceramide profile. The most obvious discrepancies were found between heat-separated epidermis and skin scrub. The reproducibility was high for scraping and skin scrub, but was lowest for heat-separated epidermis. Furthermore, this study revealed a marked influence of ceramide standards on the results regarding the ceramide profile. Scraping and skin scrub are comparably suitable methods for skin lipid sampling, whereas the analysis of heat-separated epidermis may not be the method of first choice.

Transcription Factor Ctip2 Controls Epidermal Lipid Metabolism and Regulates Expression of Genes Involved in Sphingolipid Biosynthesis during Skin Development

The stratum corneum is composed of protein-enriched corneocytes embedded in an intercellular matrix of nonpolar lipids organized as lamellar layers and give rise to epidermal permeability barrier (EPB). EPB defects play an important role in the pathophysiology of skin diseases such as eczema. The transcriptional control of skin lipid metabolism is poorly understood. We have discovered that mouse lacking a transcription factor COUP-TF interacting protein 2 (Ctip2) exhibit EPB defects including altered keratinocyte terminal differentiation, delayed skin barrier development and interrupted neutral lipid distribution in the epidermis. We adapted herein a targeted lipidomic approach using mass spectrometry, and have determined that Ctip2−/− mice (germline deletion of Ctip2 gene) display altered composition of major epidermal lipids such as ceramides and sphingomyelins compared to wildtype at different stages of skin development. Interestingly, expressions of several genes involved in skin sphingolipid biosynthesis and metabolism were altered in mutant skin. Ctip2 was found to be recruited to the promoter region of a subset of those genes, suggesting their possible direct regulation by Ctip2. Our results confirm an important role of Ctip2 in regulating skin lipid metabolism and indicate that profiling of epidermal sphingolipid could be useful for designing effective strategies to improve barrier dysfunctions.

Topical apigenin improves epidermal permeability barrier homoeostasis in normal murine skin by divergent mechanisms

The beneficial effects of certain herbal medicines on cutaneous function have been appreciated for centuries. Among these agents, chrysanthemum extract, apigenin, has been used for skin care, particularly in China, for millennia. However, the underlying mechanisms by which apigenin benefits the skin are not known. In this study, we first determined whether topical apigenin positively influences permeability barrier homoeostasis, and then the basis thereof. Hairless mice were treated topically with either 0.1% apigenin or vehicle alone twice daily for 9days. At the end of the treatments, permeability barrier function was assessed with either an electrolytic water analyzer or a Tewameter. Our results show that topical apigenin significantly enhanced permeability barrier homoeostasis after tape stripping, although basal permeability barrier function remained unchanged. Improved barrier function correlated with enhanced filaggrin expression and lamellar body production, which was paralleled by elevated mRNA levels for the epidermal ABCA12. The mRNA levels for key lipid synthetic enzymes also were upregulated by apigenin. Finally, both cathelicidin-related peptide and mouse beta-defensin 3 immunostaining were increased by apigenin. We conclude that topical apigenin improves epidermal permeability barrier function by stimulating epidermal differentiation, lipid synthesis and secretion, as well as cutaneous antimicrobial peptide production. Apigenin could be useful for the prevention and treatment of skin disorders characterized by permeability barrier dysfunction, associated with reduced filaggrin levels and impaired antimicrobial defenses, such as atopic dermatitis.

Topical Antihistamines Display Potent Anti-Inflammatory Activity Linked in Part to Enhanced Permeability Barrier Function

Systemic antagonists of the histamine type 1 and 2 receptors (H1/2r) are widely used as anti-pruritics and central sedatives, but demonstrate only modest anti-inflammatory activity. Because many inflammatory dermatoses result from defects in cutaneous barrier function, and because keratinocytes express both Hr1 and Hr2, we hypothesized that H1/2r antagonists might be more effective, if they were used topically to treat inflammatory dermatoses. Topical H1/2r antagonists additively enhanced permeability barrier homeostasis in normal mouse skin by: i) stimulation of epidermal differentiation, leading to thickened cornified envelopes; and ii) enhanced epidermal lipid synthesis and secretion. Since barrier homeostasis was enhanced to a comparable extent in mast cell-deficient mice, with no further improvement following application of topical H1/2r antagonists, H1/2r antagonists likely oppose mast cell-derived histamine. In four immunologically-diverse, murine disease models, characterized by either inflammation alone (acute irritant contact dermatitis, acute allergic contact dermatitis), or by prominent barrier abnormalities (subacute allergic contact dermatitis, atopic dermatitis), topical H1/2r agonists aggravated, while H1/2r antagonists improved inflammation and/or barrier function. The apparent ability of topical H1r/2r antagonists to target epidermal H1/2r could translate into increased efficacy in the treatment of inflammatory dermatoses, likely due to decreased inflammation and enhanced barrier function. These results could shift current paradigms of antihistamine utilization from a predominantly-systemic to a topical approach.

Design and development of ethosomal transdermal drug delivery system of valsartan with preclinical assessment in Wistar albino rats

Valsartan (VLT) is a highly selective and orally active antihypertensive drug. However, its oral administration is associated with drawbacks like low bioavailability. The objective of this study was to design and develop a transdermal delivery system for VLT using ethosomal carriers to investigate their enhanced transdermal delivery potential. VLT ethosomes were prepared by cold method. VLT ethosomes were characterized by scanning electron microscopy. The prepared ethanolic liposomes were characterized to be spherical having low polydispersity of nano-size range with good entrapment efficiency. ETC5 ethosomal suspension with 4% of phospholipon 90H and 40% of ethanol was found to have highest entrapment efficiency, i.e. 80.230 ± 0.8748%. The permeation study of ethosomes was evaluated by ex vivo diffusion study through rat abdominal skin using Franz’s diffusion cells and ETC5 ethosomal suspension was found to have highest permeation with flux of 92.819 ± 1.539 µg/cm2/h, when compared to the permeation profiles of drug solutions either in water or in a water–ethanol mixture. Transdermal application of ethosomal VLT on Wistar rats showed better and prolonged antihypertensive activity in comparison to orally administered VLT suspension by virtue of transdermal permeation through Wistar rat skin. Histopathological study of skin applied with ETC5 showed intercellular permeation across skin by dissolving intercellular lipids in epidermis without causing any rigorous changes in the skin cellular structure. In conclusion, ethosomes enabled the transdermal permeation of VLT, which amply proves its superiority over oral administration for antihypertensive treatment.

Interactions between FATP4 and ichthyin in epidermal lipid processing may provide clues to the pathogenesis of autosomal recessive congenital ichthyosis

BackgroundAutosomal recessive congenital ichthyosis (ARCI) is caused by mutations in ≥10 different genes, of which transglutaminase-1 (TGM1) predominates. A rare form is ichthyosis prematurity syndrome (IPS) caused by mutations in SLC27A4 encoding fatty acid transporter protein 4 (FATP4), believed to be an acyl-CoA synthetase activating long- and very-long-chain FA. Another ARCI is caused by mutations in NIPAL4, coding for ichthyin, which is proposed to be a magnesium transporter or a trans-membrane receptor. A possible interaction between FATP4 and ichthyin has not been studied before. ObjectiveTo find common denominators in the pathogenesis of ARCI. MethodsFATP4 and ichthyin were analyzed by immunofluorescence and proximity ligation assay (PLA) in healthy and ARCI patient skin and in in vitro models of ARCI epidermis. ResultsBoth proteins were expressed in the upper stratum granulosum of normal epidermis and PLA confirmed a close interaction between FATP4 and ichthyin. In IPS skin lacking FATP4 we found reduced ichthyin expression and this finding could be reproduced in organotypic epidermis with siRNA silenced SLC27A4. In contrast, increased FATP4 staining was found in patients with ichthyin (NIPAL4) mutations and in organotypic epidermis with silenced NIPAL4. In patients with TGM1 mutations, the expression of both FATP4 and ichthyin was increased, but the PLA signal was low probably indicating a malfunctioning protein interaction. ConclusionOur study suggests that FATP4, ichthyin and TGM1 interact in lipid processing essential for maintaining the epidermal barrier function. It is also hypothesized that ichthyin serves as Mg2+-transporter for FATP4 in this process.

Urea Uptake Enhances Barrier Function and Antimicrobial Defense in Humans by Regulating Epidermal Gene Expression

Urea is an endogenous metabolite, known to enhance stratum corneum hydration. Yet, topical urea anecdotally also improves permeability barrier function, and it appears to exhibit antimicrobial activity. Hence, we hypothesized that urea is not merely a passive metabolite, but a small-molecule regulator of epidermal structure and function. In 21 human volunteers, topical urea improved barrier function in parallel with enhanced antimicrobial peptide (LL-37 and β-defensin-2) expression. Urea both stimulates expression of, and is transported into keratinocytes by two urea transporters, UT-A1 and UT-A2, and by aquaporin 3, 7 and 9. Inhibitors of these urea transporters block the downstream biological effects of urea, which include increased mRNA and protein levels for: (i) transglutaminase-1, involucrin, loricrin and filaggrin; (ii) epidermal lipid synthetic enzymes, and (iii) cathelicidin/LL-37 and β-defensin-2. Finally, we explored the potential clinical utility of urea, showing that topical urea applications normalized both barrier function and antimicrobial peptide expression in a murine model of atopic dermatitis (AD). Together, these results show that urea is a small-molecule regulator of epidermal permeability barrier function and antimicrobial peptide expression after transporter uptake, followed by gene regulatory activity in normal epidermis, with potential therapeutic applications in diseased skin.

Skin Ceramide Alterations in First-Episode Schizophrenia Indicate Abnormal Sphingolipid Metabolism

There is considerable evidence for specific pathology of lipid metabolism in schizophrenia, affecting polyunsaturated fatty acids and in particular sphingolipids. These deficits are assumed to interfere with neuronal membrane functioning and the development and maintenance of myelin sheaths. Recent studies suggest that some of these lipid pathologies might also be detected in peripheral skin tests. In this study, we examined different skin lipids and their relation to schizophrenia. We assessed epidermal lipid profiles in 22 first-episode antipsychotic-naïve schizophrenia patients and 22 healthy controls matched for age and gender using a hexan/ethanol extraction technique and combined high-performance thin-layer chromatography/gas-chromatography. We found highly significant increase of ceramide AH and NH/AS classes in patients and decrease of EOS and NP ceramide classes. This is the first demonstration of specific peripheral sphingolipid alterations in schizophrenia. The results support recent models of systemic lipid pathology and in particular of specific sphingolipids, which are crucial in neuronal membrane integrity. Given recent findings showing amelioration of psychopathology using fatty acid supplementation, our findings also bear relevance for sphingolipids as potential biomarkers of the disease.

Structure and function of the epidermis related to barrier properties

The most important function of the skin is the formation of a barrier between the “inside” and the “outside” of the organism, which prevents invasion of pathogens and fends off chemical assaults as well as the unregulated loss of water and solutes. The physical barrier is mainly localized in the stratum corneum, which consists of protein-enriched cells and lipid-enriched intercellular domains. Any modifications in epidermal differentiation and lipid composition results in altered barrier function, a central event in various skin alterations and diseases. This contribution presents a brief description of the structure of the skin, paying attention to the most important components responsible for skin barrier function.

Nrf2 links epidermal barrier function with antioxidant defense

The skin provides an efficient permeability barrier and protects from microbial invasion and oxidative stress. Here, we show that these essential functions are linked through the Nrf2 transcription factor. To test the hypothesis that activation of Nrf2 provides skin protection under stress conditions, we determined the consequences of pharmacological or genetic activation of Nrf2 in keratinocytes. Surprisingly, mice with enhanced Nrf2 activity in keratinocytes developed epidermal thickening, hyperkeratosis and inflammation resembling lamellar ichthyosis. This resulted from upregulation of the cornified envelope proteins small proline-rich proteins (Sprr) 2d and 2h and of secretory leukocyte peptidase inhibitor (Slpi), which we identified as novel Nrf2 targets in keratinocytes. Since Sprrs are potent scavengers of reactive oxygen species and since Slpi has antimicrobial activities, their upregulation contributes to Nrf2's protective function. However, it also caused corneocyte fragility and impaired desquamation, followed by alterations in the epidermal lipid barrier, inflammation and overexpression of mitogens that induced keratinocyte hyperproliferation. These results identify an unexpected role of Nrf2 in epidermal barrier function, which needs to be considered for pharmacological use of Nrf2 activators.

Ceramide profile in hypohidrotic ectodermal dysplasia

Hypohidrotic ectodermal dysplasia (HED) is a rare genetic disease. The clinical presentation includes lack of sweating ability, and an often widely spread dermatitis resembling atopic dermatitis (AD). In AD, the skin-barrier defect is partly ascribed to the altered lipid profile in the stratum corneum and partly to mutations of the filaggrin genes. To our knowledge, no data are available about the epidermal lipid profile of HED. To compare the ceramide profile for patients with HED and AD. The ceramide profile and ceramide/cholesterol ratio were compared between patients with HED (n = 7) and patients with AD (n = 21), using cyanoacrylate to take biopsy samples from the stratum corneum. Lipids were extracted from the biopsies and analysed using high-performance thin-layer chromatography. The lipid profiles of HED and AD were similar in distribution, apart from ceramide 1, which was significantly higher in HED (P = 0.04). The increased ceramide 1 level found in HED compared with AD is known to play a role in the structure of the lipid bilayers. However, further studies are needed to identify the functional significance of these observations and thereby elucidate differences in the skin barrier between HED and AD.

TCF/Lef1-Mediated Control of Lipid Metabolism Regulates Skin Barrier Function

Defects in the function of the skin barrier are associated with a wide variety of skin diseases, many of which are not well characterized at the molecular level. Using Lef1 (lymphoid enhancer-binding factor 1) dominant-negative mutant mice, we demonstrate here that altered epidermal TCF (T cell factor)/Lef1 signaling results in severe impairment of the stratum corneum skin barrier and early postnatal death. Barrier defects were accompanied by major changes in lipid composition and ultrastructural abnormalities in assembly and extrusion of lipid lamellae of the interfollicular epidermis, as well as abnormal processing of profilaggrin. In contrast, tight-junction formation and stratified organization of the interfollicular epidermis was not obviously disturbed in Lef1 mutant mice. Molecular analysis revealed that TCF/Lef1 signaling regulates expression of lipid-modifying enzymes, such as Elovl3 and stearoyl coenzyme A desaturase 1 (SCD1), which are key regulators of barrier function. Promoter analysis and chromatin immunoprecipitation experiments indeed showed that SCD1 is a direct target of Lef1. Together, our data demonstrate that functional TCF/Lef1 signaling governs important aspects of epidermal differentiation and lipid metabolism, thereby regulating skin barrier function.

Ichthyin/NIPAL4 localizes to keratins and desmosomes in epidermis and Ichthyin mutations affect epidermal lipid metabolism

Autosomal recessive congenital ichthyosis (ARCI) is a group of disorders characterized by abnormal desquamation of the skin and a disrupted epidermal water barrier. Ichthyin/NIPAL4 gene mutations have been identified in a subgroup of ARCI patients, but the role of ichthyin in epidermis remains elusive. In order to obtain new insights concerning the characteristics of ichthyin and the ARCI pathogenesis, we studied the expression and localization of ichthyin and related epidermal components in cultured keratinocytes and skin sections from patients with Ichthyin mutations and healthy controls. We observed an up-regulation of Ichthyin mRNA levels after in vitro differentiation of keratinocytes from both a patient with Ichthyin mutations and controls. Confocal and electron microscopy analyses of immunolabeled skin sections revealed that ichthyin localizes to desmosomes and keratins in both patients with mutant Ichthyin and controls, with an increased immunolabeling in patients. Nile red lipid analysis of skin sections exposed intra-cellular lipid accumulations in cells of the granular and cornified layers in patients but not in controls, consistent with the pathognomonic lipid membrane structures previously identified in epidermis from patients. Our combined findings indicate that ichthyin is associated with keratins and desmosomes in epidermis and is involved in lipid metabolism, possibly through processing of lamellar bodies. These results provide new clues to the understanding of the epidermal water barrier and the pathogenesis in ARCI.

PNPLA1 mutations cause autosomal recessive congenital ichthyosis in golden retriever dogs and humans

Ichthyoses comprise a heterogeneous group of genodermatoses characterized by abnormal desquamation over the whole body, for which the genetic causes of several human forms remain unknown. We used a spontaneous dog model in the golden retriever breed, which is affected by a lamellar ichthyosis resembling human autosomal recessive congenital ichthyoses (ARCI), to carry out a genome-wide association study. We identified a homozygous insertion-deletion (indel) mutation in PNPLA1 that leads to a premature stop codon in all affected golden retriever dogs. We subsequently found one missense and one nonsense mutation in the catalytic domain of human PNPLA1 in six individuals with ARCI from two families. Further experiments highlighted the importance of PNPLA1 in the formation of the epidermal lipid barrier. This study identifies a new gene involved in human ichthyoses and provides insights into the localization and function of this yet uncharacterized member of the PNPLA protein family.

Morphology of Rathke's Glands in the Snapping Turtle, Chelydra serpentina, with Comments on the Presence of Multilaminar Lamellar Bodies in Turtles

I examined the histology and ultrastructure of Rathke’s glands in hatchling, juvenile, and adult snapping turtles (Chelydra serpentina). This species possesses four pairs of Rathke’s glands (i.e., one axillary and three inframarginals) that are embedded beneath marginal bones and are named primarily according to the anatomical location of their orifices. These integumentary glands are similar anatomically and ultrastructurally to one another. Each gland is comprised of a single, highly vascularized, secretory lobule, which is surrounded by a thick tunic of asymmetrically arranged, striated muscle. Two types of large secretory vacuoles characterize most of the holocrine cells produced by a relatively thin secretory epithelium. My results support previous studies that suggest that the chief secretory material of the mostly larger, dark-staining Type 1 secretory vacuole is a glycoprotein complex. The mostly translucent Type 2 secretory vacuole may contain osmiophilic granules that exhibit variously sized lamellar bodies, whose multilaminar structural design is reminiscent of an epidermal lipid delivery system in many vertebrates. The functional role of Rathke’s gland secretions in Chelydra serpentina and in other turtles remains unknown.