An ex vivo RNA trans-splicing strategy to correct human generalized severe epidermolysis bullosa simplex.

BACKGROUND: Generalized severe epidermolysis bullosa simplex (EBS-gen sev) is a genetic blistering skin disease in which autosomal dominant mutations in either the keratin KRT5 or KRT14 genes lead to impaired function of the intermediate filament cytoskeleton in the basal epidermis. Here we present an ex vivo RNA trans-splicing-based therapeutic approach to correct the phenotype. OBJECTIVES: To correct a mutation within exon 1 of the KRT14 gene, using a 5'-trans-splicing approach, where any mutation within the first seven exons could be replaced by a single therapeutic molecule. METHODS: A therapeutic RNA trans-splicing molecule containing wild-type exons 1-7 was stably transduced into an EBS patient-derived keratinocyte line. Trans-splicing was confirmed via reverse-transcriptase polymerase chain reaction, Western blotting and immunofluorescence microscopy. Skin equivalents generated from corrected keratinocytes were grafted onto nude mice and analysed about 8 weeks post-transplantation for regular epidermal stratification, trans-splicing-induced green fluorescent protein expression and blistering. RESULTS: Transplanted skin equivalents generated from trans-splicing-corrected patient keratinocytes showed a stable and blister-free epidermis. KRT14 correction disrupted EBS-gen sev-associated proinflammatory signalling, as shown at the mRNA and protein levels. Disruption of the pathogenic feedback loop in addition to overall downregulation of KRT14 expression highlighted the effect of KRT14 correction on the EBS pathomechanism. CONCLUSIONS: Our data demonstrate that trans-splicing-mediated mRNA therapy is an effective method for the correction of dominantly inherited KRT14 mutations at the transcriptional level. This results in the rescue of the EBS-gen sev phenotype and stabilization of the epidermis in a xenograft mouse model.

Construction and validation of an RNA trans-splicing molecule suitable to repair a large number of COL7A1 mutations.

RNA trans-splicing has become a versatile tool in the gene therapy of monogenetic diseases. This technique is especially valuable for the correction of mutations in large genes such as COL7A1, which underlie the dystrophic subtype of the skin blistering disease epidermolysis bullosa. Over 800 mutations spanning the entire length of the COL7A1 gene have been associated with defects in type VII collagen, leading to excessive fragility of epithelial tissues, the hallmark of dystrophic epidermolysis bullosa (DEB). In the present study, we designed an RNA trans-splicing molecule (RTM) that is capable of repairing any given mutation within a 4200 nucleotide region spanning the 3' half of COL7A1. The selected RTM, RTM28, was able to induce accurate trans-splicing into endogenous COL7A1 pre-mRNA transcripts in a type VII collagen-deficient DEB patient-derived cell line. Correct trans-splicing was detected at the RNA level by semiquantitative RT-PCR and correction of full-length type VII collagen was confirmed at the protein level by immunofluorescence and western blot analyses. Our results demonstrate that RTM28, which covers >60% of all mutations reported in DEB and is thus the longest RTM described so far for the repair of COL7A1, represents a promising candidate for therapeutic applications.

A population based evaluation of the mode of delivery in association with infertility treatment from 1990-2012.

BACKGROUND: This study refers to population based data and investigates the development of the mode of delivery associated with infertility treatment over the last 23 years. METHODS: All 1 202,557 deliveries in Hesse, Germany, between 1990 and 2012 were assessed. 2.2% of the study population, 26,761, had a delivery subsequent to infertility treatment based on the Hessian Perinatal Registry (HEPE). An evaluation in this subgroup was performed investigating the associations between the mode of delivery and the gestational week and the mother's age. RESULTS: A continuous and significant (p<0.01) increase of cesarean section (CS) rates subsequent to infertility treatment (1990: 41,3%; 2012: 55,9%) as well as a conversely also significant (p<0.01) reduction of vaginal operative and spontaneous deliveries associated with infertility treatment between 1990 and 2012 was found. Furthermore, the preterm delivery rate and the proportion of deliveries of parturients older than 35 years of age in association with infertility treatment raised over the last years. Rates of full-term deliveries and deliveries of women younger than 35 years remained stable during the observation period. DISCUSSION: The rate of cesarean section is continuously rising over the last 23 years with regard to parturients subsequent to infertility treatment. The CS rate is significantly higher compared to women with a spontaneous pregnancy and in comparison to the data from 20 years ago. Most recently, the number of CS (51,2%) exceeded the number of vaginal deliveries (48,8%) in Hesse subsequent to infertility treatment for the first time.

Trends of vaginal birth after cesarean delivery in Germany from 1990 to 2012: a population-based study.

BACKGROUND: The increasing incidence of cesarean deliveries (CD) in the western world is consequently leading to a rising number of antenatal counselling of pregnant women with a history of previous CD. To counteract the increasing trend of cesarean deliveries, the concept of vaginal birth after cesarean delivery (VBAC) may represent an alternative. The aim of the present study was to longitudinally investigate the incidence of VBAC and compare the changes within all deliveries during 23 years of follow-up. METHODS: In this study we analyzed data from 1 202 557 deliveries in Hesse, Germany from 1990 to 2012. In total, 131 629 births have been identified to have at least one CD in the patients' medical history. We grouped the patients into 3 categories: vaginal spontaneous birth subsequent to CD, vaginal-operative birth subsequent to CD and repeated CD. RESULTS: After previous CD, 32.1% of the patients delivered spontaneously, 4.0% delivered vaginal-operative and 63.8% had a repeated CD. The rates changed from 40.4, 7.5 and 52.1% in the year 1990 to 23.3, 2.8 and 73.9% in the year 2012 for vaginal spontaneous births, vaginal-operative births and for repeated CDs, respectively (p<0.01). We noticed a decline of 17.1 and 4.7% in spontaneous births after Cesarean and vaginal operative births respectively during the observation period. Notably, we report a dramatic increase of 21.8% of repeated CDs during the past 23 years (p<0.01). With regard to the non-affected group including all deliveries, we observed a decrease of 17% in spontaneous deliveries from 1990 to 2012 (75.9 vs. 58.9%). Vaginal operative delivery rates changed from 6.9% in 1990 to 5.9% in 2012. Consequently, CD rates increased from 17.2% in 1990 to 35.2% in the year 2012 (p<0.01). The differences between all 3 subgroups were significantly different (p<0.001). DISCUSSION: Cesarean rates in Germany have reached an all-time high, while VBAC follows a continuous decrease. The current rate of VBAC is almost the half of that in the year 1990 (26.1 vs. 47.9%). Promotion of a trial of labor (TOL) after low transverse CD in those women who desire 3 or more children may increase the VBAC success rates and reduce maternal morbidity.

A Population-based study on trends in the mode of delivery subsequent to stillbirth, infant mortality and severe infant malformations in the state of Hesse, Germany between 1990 and 2012.

BACKGROUND: Women giving birth to a child with severe infant malformations or suffering a stillbirth certainly experience a mental trauma. Therefore the objective of the present study is to examine the incidence of deliveries subsequent to such a traumatic experience as well as the mode of delivery. Secondly, the results are compared to the changes of a non-affected group of deliveries over the last 23 years in the state of Hesse, Germany. METHODS: The total obstetric dataset of the Hessian Perinatal Registry (HEPE) was assessed for women with regard to one item of the pregnancy risk factors. This particular HEPE item comprises information on the rates of stillbirth, early and late infant mortality (≤7 days,<1 year) as well as severe infantile malformations in women giving birth subsequent to such an extensive traumatic experience. The identified women were categorized with respect to the mode of delivery (spontaneous, vaginal operative or Cesarean section), pre- and full-term birth and according to 4 time-frames between 1990 and 2012. The results of women with a positive HEPE item were compared to those of women without such a traumatic experience (non-affected group) of the HEPE analyzed in the same categories and time-frames. RESULTS: The obstetric dataset from 1990 until 2012 of the HEPE comprised altogether 1 224 760 deliveries including a group of 19 726 (1.61%) deliveries subsequent to a positive result for the analyzed HEPE item. Over the duration of follow-up the rate of subsequent pregnancies following such a traumatic experience showed a significantly decrease of 43% in comparison to the previous time-frame, respectively (1990-1996: 30.3%; 2008-2013: 17%; P≤0.0001). With respect to the mode of delivery (spontaneous, vaginal operative, Cesarean section) the results in the group subsequent to the positive HEPE item (1 862, 55.5%; 70, 2.1%; 1 416, 42.2%) were significantly different (P<0.001) in comparison to the non-affected group (142 846, 59.9%; 13 875, 5.8%; 81 089, 34.0%), respectively. DISCUSSION: Rising rates of CSs and conversely decreased numbers of vaginal deliveries in most cases of full-term pregnancies contrast with the reduction of the positive HEPE item over the last 23 years. There seems to be an urgent need for action since this observed critical trend remains highly significant in comparison with the rising trend of CS in general. Pre-pregnancy counseling and continuous monitoring during pregnancy and delivery in consciousness of evident risk factors may be an opportunity to cope with these serious findings.

Formation of deposits on the surfaces of superheaters and economisers of MSW incinerator plants.

Mineralogical and chemical investigations of deposits from superheaters and economisers from a MSWI plant in Mannheim, Germany, lead to a classification system which provides information about the most critical parameters leading to fouling and corrosion. With the help of this classification system parameters like the geometry of boilers and the waste input can be changed in order to prolong run times between revisions and enhance energy efficiency of MSWI plants.

Effect of short-term natural weathering on MSWI and wood waste bottom ash leaching behaviour.

Short term natural weathering was applied on municipal solid waste (MSW) and wood waste incinerator bottom ash (BA). The materials were analysed at different steps of treatment and characterized for chemical and mineralogical composition. Both short and long term leaching behaviour of main elements and heavy metals were investigated as well. Lead, zinc and copper were the main heavy metals to be released. After 12 weeks of treatment the concentration of leached zinc decreased. Lead concentration was not found to be influenced by pH and decreased only for the biomass samples. Weathering did not have beneficial effects on copper leaching, which was well described by complexation processes with DOC. The findings from the experimental campaign indicated that weathering reactions improved the mineral stability of the analysed materials but, in contrast with previous works, the treatment was not sufficient to guarantee pH stability and to comply with leaching law limits.

[Amniotic membrane transplantation in proven ulcerative herpetic keratitis: successful anti-inflammatory treatment in time]. / Amnionmembrantransplantation bei gesicherter ulzerativer Herpeskeratitis: Erfolgreiche Entzündungshemmung zum richtigen Zeitpunkt.

BACKGROUND: In active herpetic keratitis, treatment with systemic and topical aciclovir together with low dosed-out steroids has been successful for few decades. However, in cases with severe herpetic ulcer and melting inflammation, emergency penetrating keratoplasty is necessary, which carries high risk for recurrent herpetic keratitis and rejection. The anti-inflammatory effect of amniotic membrane transplantation can be used as a first surgical step in order to gain time for planned penetrating keratoplasty. The results are analysed in this retrospective case series. PATIENTS: Twelve eyes of 12 patients with clinically highly active herpetic ulcer without healing despite topical and systemic aciclovir were treated with amniotic membrane transplantation. Ten of 12 eyes (83 %) showed positive PCR testing for HSV-1. In one case, the molecular genetic result was negative, one analysis for HSV was not done. For surgery, debridement of the margins and the ground of the ulcer was done with steam heated cauterization. Afterwards, an amniotic membrane was made to cover the defect and fixed with single sutures. Peri- and postoperatively, the patients were treated with systemic aciclovir (4000 mg daily one week, 2000 mg daily 2 weeks) and gentamicin ointment topically. RESULTS: Mean follow-up was 6.8 months (1-12.5). After 25 days in the mean (11-34), nine of 12 patients experienced an intact epithelial corneal surface together with a clinically clear reduction of the inflammation. Three other patients did not recover and were treated with emergency penetrating keratoplasty after 31 days in mean (25-35). Visual acuity did not improve after amniotic membrane transplantation except in one case. CONCLUSION: The known anti-inflammatory potential of amniotic membrane causes a reduction of the inflammatory process in herpetic ulcer. A precondition is a sufficient antiherpetic and anti-inflammatory therapy and a decision for amniotic membrane transplantation in time in order to avoid an emergency penetrating keratoplasty. This gives the opportunity to perform planned penetrating keratoplasty with optimal HLA-match and postoperative immunosuppression.

[Acquired esotropia at advanced age]. / Erworbenes Innenschielen im Alter.

BACKGROUND: Patients, most of whom are in their 8th decade of life, can suffer from a minimal acquired esotropia. They complain of blurred vision instead of diplopia due to their confusion. This is accompanied with the risk that the investigation covers only monocular disturbances. PATIENTS AND METHODS: Of 39 such patients, 22 underwent surgery while 10 were fitted with prisms. Another 3 recovered spontaneously and became, therefore, classified as VIth nerve palsies although no limitation of the abduction could be stated. Four patients refused any therapy, and 2 of them recovered in the course of many years. RESULTS: After 2 to 7 years, the individuals corrected with prisms were still compensated while within the operated cohort 3 had a recurrence after years. CONCLUSIONS: In almost all cases the prescription of separate glasses for distance with about 3 Delta base out on either side compensates the deviation for several years. In surgical units one finds a summation of chronically decompensated patients with obvious diplopia who need a bimedial recession. Lateral rectus resections are not appropriate as they lead to actual overabduction and final recurrence as well.

DNA methylation changes in multiple myeloma.

Using a candidate gene approach, we analyzed the methylation status of the promoter-associated CpG islands of 11 well-characterized tumor suppressor genes by methylation-specific polymerase chain reaction in five multiple myeloma (MM) cell lines and 56 patients with malignant plasma cell disorders. The frequency of aberrant methylation among the patient samples was 46.4% for SOCS-1, 35.7% for p16, 21.4% for E-cadherin, 12.5% for DAP kinase and p73, 1.8% for p15, MGMT as well as RARbeta, and 0% for TIMP-3, RASSF1A and hMLH1. We found at least one hypermethylated gene in 80.4% of the primary patient samples, while 33.9% harbored two or more hypermethylated genes. For the first time, we show that p73 may be hypermethylated in MM and thus be involved in the pathogenesis of plasma cell disorders. Hypermethylation of p16 at diagnosis was associated with a poorer prognosis. In patients with plasma cell leukemia, we found frequent simultaneous hypermethylation of p16, E-cadherin and DAP kinase. We conclude that aberrant methylation of tumor suppressor genes is a common event in malignant plasma cell disorders and that there is a correlation between methylation patterns and clinical characteristics in MM patients.

[Impairment of safety in navigation caused by alcohol: impact on visual function]. / Beeinträchtigung der Sicherheit im Schiffsverkehr durch Alkohol. Einfluss auf das Visuelle System.

BACKGROUND: So far in Germany, no legally binding standards for blood alcohol concentration exist that prove an impairment of navigability. The aim of our interdisciplinary project was to obtain data in order to identify critical blood alcohol limits. In this context the visual system seems to be of decisive importance. MATERIALS AND METHODS: 21 professional skippers underwent realistic navigational demands soberly and alcoholized in a sea traffic simulator. The following parameters were considered: visual acuity, stereopsis, color vision, and accommodation. RESULTS: Under the influence of alcohol (average blood alcohol concentration: 1.08 per thousand ) each skipper considered himself to be completely capable of navigating. While simulations were running, all of the skippers made nautical mistakes or underestimated dangerous situations. Severe impairment in visual acuity or binocular function were not observed. Accommodation decreased by an average of 18% ( p=0.0001). In the test of color vision skippers made more mistakes ( p=0.017) and the time needed for this test was prolonged ( p=0.004). CONCLUSIONS: Changes in visual function as well as vegetative and psychological reactions could be the cause of mistakes and alcohol should therefore be regarded as a severe risk factor for security in sea navigation.

[Impairment of safety in navigation caused by alcohol: pupillography and measurement of pupil movements]. / Beeinträchtigung der Sicherheit im Schiffsverkehr durch Alkohol. Pupillographie und Augenbewegungsmessungen.

OBJECTIVE: To evaluate data in an interdisciplinary project (ophthalmology, forensic medicine, internal medicine, psychology,and nautical science) in order to identify critical blood alcohol limits in sea navigation. METHODS: A sea traffic simulator was employed for realistic nautical demands on 21 professional experienced skippers under sober and alcoholized conditions (target blood alcohol concentration: 1.0 per thousand ). After simulated navigation, pupil light reflex, spontaneous pupil movements,nystagmus,and saccades were evaluated by pupillography. Modification of the pupillograph enabled us also to measure optokinetic nystagmus. RESULTS: Evaluation of the pupil light reflex revealed obvious changes in the extent of relative contraction and in redilatation time under the influence of alcohol. Diminished vigilance could be observed in all of the skippers when optokinetic nystagmus was tested. CONCLUSION: The pupillograph represents a suitable device for measuring functions of the visual and vegetative systems. Thus, the impact of these functions on nautical capability can be demonstrated. If further investigations such as ophthalmological, medical, psychological, and nautical evaluations are taken into account, it could be determined that blood alcohol levels of 1.0 per thousand may exclude safe navigation.

Keratins: a structural scaffold with emerging functions.

Intermediate filament proteins form an essential part of the cytoskeleton and provide topological order to cells and tissues. These features result from their intrinsic property of self-organization and their response to extrinsic cues. Keratins represent the largest subgroup among all intermediate filament proteins and are differentially expressed as pairs of type I and type II intermediate filament proteins in epithelia. Their primary function is to impart mechanical strength to cells. This function is illustrated by patients with keratin mutations and by gene-deficient mice. Additional functions include their participation in the response to stress, cell signalling and apoptosis, and thus the keratin cytoskeleton appears far more dynamic than previously anticipated. This may result from hyperphosphorylation and possibly from interaction with associated proteins. How signalling networks affect keratin organization, turnover and function and vice versa will be a major challenge for future investigations.

[Pars plana vitrectomy in cystoid macular edema of different forms of chronic uveitis]. / Pars-plana-Vitrektomie bei zystoidem Makulaödem bei verschiedenen Formen der chronischen Uveitis.

BACKGROUND: Cystoid macular edema (CME) is a common complication in different forms of chronic uveitis. In spite of immunosuppressive and anti-inflammatory therapy, chronic or relapsing courses can occur which may have a negative impact on visual prognosis. Pars plana vitrectomy (PPV) is known to positively influence chronic uveitis. This retrospective study was performed to investigate the role of PPV in the therapy refractive uveitic CME. PATIENTS AND METHODS: PPV for CME was performed in eyes with CME in intermediate uveitis (IMU, n=42), chronic iridocyclitis in juvenile rheumatoid arthritis (CIC, n=14) and multifocal chorioretinitis (MFC, n=12). In none of the eyes had immunosuppressive and/or anti-inflammatory therapy or anti-edema treatment (e.g.acetazolamide) led to regression of the CME. After a postoperative follow-up period of 7 and 106 months all patients were re-examined. RESULTS: Postoperative complete or partial regression of CME was observed as follows: IMU: 25/42 (59.5%), CIC: 8/14 (57.1%),MFC: 5/12 (41.7%). A significant increase in visual acuity of 2 lines and more was observed in 50%,71.4% and 41.7% of eyes, respectively. In the long-term follow-up best functional results were achieved in eyes with IMU. CONCLUSIONS: Response to PPV was variable according to the type of underlying form of uveitis. The lowest success rate could be observed in eyes with MFC. Although the postoperative regression rate of CME was satisfactory in eyes with CIC, long-term visual acuity results were disappointing due to secondary complications of CIC in this young age group. Best results were achieved in patients with IMU (statistically not significant). A multicenter study in a larger series of patients is needed to investigate the exact role of PPV in different forms of chronic uveitis.

[Alcohol-induced morphologic and biochemical corneal changes]. / Alkoholinduzierte morphologische und biochemische Hornhautveränderungen.

In our cornea bank, it was noticed that corneas from donors with alcoholism seemed to be of lower quality than corneas from other donors. High blood ethanol concentrations can induce high alcohol concentrations in aqueous and vitreous humor. This could be demonstrated in the case of a lethal alcohol intoxication. We conducted in vitro experiments to clarify the question of alcohol-induced changes of corneas. The corneas were stored in a standardized culture medium including ethanol, formic acid, methanol, and acetaldehyde in concentrations to be expected in chronic alcoholism. During cultivation over 4 weeks, endothelial morphology and extent of aspartic acid racemization in stromal proteins were evaluated. The extent of aspartic acid racemization served as a biochemical parameter of alcohol-induced protein changes of the corneas. In the in vitro experiments, a drop in the endothelial cell counts could be seen in corneas stored in culture medium with acetaldehyde. The extent of aspartic acid racemization increases in corneas stored in medium containing ethanol, which obviously is the result of alcohol-induced protein degradation. High concentrations of ethanol and its metabolics can be detected in vitreous and aqueous humor in chronic alcoholism. Clearly, these concentrations have a direct toxic effect on the corneal endothelium (acetaldehyde) and on the stromal proteins (ethanol).

Pars-plana vitrectomy in cystoid macular edema associated with intermediate uveitis.

BACKGROUND: Cystoid macular edema (CME) is a common complication in the course of intermediate uveitis. In spite of systemic therapy with steroids or carbonic anhydrase inhibitors, persistence of CME is observed. Pars plana vitrectomy (PPV) is known to influence the course of intermediate uveitis positively. The present study was performed to investigate the role of PPV in the therapy of CME in intermediate uveitis. MATERIALS AND METHODS: Forty-two eyes of 32 patients were re-examined after PPV for CME. In all eyes fluorescein angiography was performed. Average age at the time of surgery was 31.9 years (range 6-64 years). All patients had received systemic corticosteroid and/or immunosuppressive treatment during the course of their disease. In some patients systemic therapy with carbonic anhydrase inhibitors was performed. The mean duration of postoperative follow-up was 20.2 months (range 6-102 months). RESULTS: Preoperative visual acuity (VA) in all eyes was between 1/10 and 0.5. Total regression of CME after surgery was observed in 18 of 42 eyes (42.8%), partial improvement in 7 eyes (16.7%). In 13 of 42 eyes (30.9%) the CME remained unchanged. Twenty-one of 42 eyes (50.0%) experienced a postoperative improvement of VA of 2 lines or more. In 18 of 42 eyes (42.8%) there was no change; in 3 eyes (7.2%) VA was less. In the long-term follow-up the corresponding results were slightly worse (17/17/8 eyes) due to secondary complications. In the majority of patients systemic medical therapy could be reduced or discontinued. CONCLUSION: Pars-plana vitrectomy led to regression of CME in 59% of cases and to subsequent improvement of VA in 50% of eyes with intermediate uveitis. PPV should be considered soon after medical therapy has been shown to be ineffective.

Energetic stress induces premature aging of diploid human fibroblasts (Wi-38) in vitro.

Oxidative phosphorylation is the main endogenous source for the generation of reactive oxygen species (ROS). In order to investigate the influence of enhanced ROS production on the in vitro senescence of Wi-38 fibroblasts, cells were cultivated in medium with elevated (hypertonic) NaCl concentrations. The number of active Na(+)/K(+)-ATPase molecules per cell was found to be increased. A rise in both respiration and glycolysis as evidenced by the increases in oxygen and glucose consumption and lactate production was revealed. Cells stayed alive in medium with NaCl concentrations of up to 0.30 M and could be adapted to growth under these hypertonic conditions (high-NaCl tolerant cells). These cells exhibited an increased cell size and protein content. A growing number of cells showed stress fibers and granulation. The proliferation rate and the maximum number of cumulative population doublings of these high-NaCl tolerant cultures were reduced and saturation density was decreased. Thus, these cells under energetic stress due to increased energy requirements for active ion transport expressed features typical for aging in vitro. We conclude therefore that energetic stress induces premature aging in human diploid fibroblasts.

Formation of a normal epidermis supported by increased stability of keratins 5 and 14 in keratin 10 null mice.

The expression of distinct keratin pairs during epidermal differentiation is assumed to fulfill specific and essential cytoskeletal functions. This is supported by a great variety of genodermatoses exhibiting tissue fragility because of keratin mutations. Here, we show that the loss of K10, the most prominent epidermal protein, allowed the formation of a normal epidermis in neonatal mice without signs of fragility or wound-healing response. However, there were profound changes in the composition of suprabasal keratin filaments. K5/14 persisted suprabasally at elevated protein levels, whereas their mRNAs remained restricted to the basal keratinocytes. This indicated a novel mechanism regulating keratin turnover. Moreover, the amount of K1 was reduced. In the absence of its natural partner we observed the formation of a minor amount of novel K1/14/15 filaments as revealed by immunogold electron microscopy. We suggest that these changes maintained epidermal integrity. Furthermore, suprabasal keratinocytes contained larger keratohyalin granules similar to our previous K10T mice. A comparison of profilaggrin processing in K10T and K10(-/-) mice revealed an accumulation of filaggrin precursors in the former but not in the latter, suggesting a requirement of intact keratin filaments for the processing. The mild phenotype of K10(-/-) mice suggests that there is a considerable redundancy in the keratin gene family.