Multiple miliary osteoma cutis is a distinct disease entity: four case reports and review of the literature.

BACKGROUND: Multiple miliary osteoma cutis (MMOC) is a rare nodular skin disease characterized by tiny bone nodules which usually form on the facial skin, typically in middle age. The aetiology of this phenomenon is poorly understood. OBJECTIVES: To search for possible bone formation progenitors and to look for a possible association with mutations in the GNAS gene (encoding the G-protein α-stimulatory subunit) and related hormonal parameters in patients with MMOC. We also reviewed the literature and discuss the aetiology and pathogenesis of adult-onset primary osteomas. METHODS: We report four cases of MMOC. Histological samples were analysed for bone morphogenetic protein (BMP)-2, BMP-4 and oestrogen receptor-α known to be involved in bone formation. Endocrinological laboratory investigations and hand X-rays were performed to exclude a systemic disease. The GNAS gene was sequenced from DNA extracted from peripheral blood in all four patients and from a skin sample in one patient to exclude somatic mutations. RESULTS: Histological analyses revealed intramembranous cutaneous bone formation resembling the findings seen in GNAS gene-based osteoma cutis disorders. However, we did not find any germline or somatic GNAS gene mutations in our patients and all laboratory investigations gave normal results. BMP-2 and -4 were expressed normally in MMOC samples, but oestrogen receptor-α was not expressed. Altogether 47 MMOC cases, 41 female and six male, have been published between 1928 and 2009. Of these cases, 55% had a history of pre-existing acne and only 15% had extrafacial osteomas. CONCLUSIONS: MMOC is a rare but distinct disease entity of unknown aetiology. Histologically, the tiny nodular osteomas show intramembranous superficial ossification but the aetiology appears to be different from GNAS-related disorders. The osteomas seem to increase slowly in number after appearing in middle age.

Increased expression of glucocorticoid receptor beta in lymphocytes of patients with severe atopic dermatitis unresponsive to topical corticosteroid.

BACKGROUND: Variable response to topical glucocorticoid therapy occurs in the treatment of severe atopic dermatitis (AD). Glucocorticoid receptor (GR)-beta does not bind glucocorticoids but antagonizes the activity of the classic GRalpha, and could thus account for glucocorticoid insensitivity. OBJECTIVES: To investigate GRalpha and GRbeta mRNA and protein expression in lymphocytes of patients with AD before and after treatment with topical corticosteroids. METHODS: Blood was collected from 11 healthy donors, 10 patients with mild AD and 13 patients with severe AD. mRNA was isolated from peripheral blood mononuclear cells. Expression of GRalpha and GRbeta mRNA was determined by reverse transcriptase-polymerase chain reaction and quantitated. Expression of the GRs was confirmed by Western blot analysis. RESULT: The expression of GRalpha mRNA was detected in all subjects. GRbeta mRNA was detected in four out of 11 healthy volunteers, five out of 10 patients with mild AD and 11 out of 13 patients with severe AD. The incidence of GRbeta mRNA expression was higher in patients with severe AD (85%) than in patients with mild AD (50%), and significantly higher than in healthy volunteers (36%, P = 0.033). Four of the 13 patients with severe AD showed a 3.3-13.2-fold increase in the expression of GRbeta mRNA during a 2-week treatment with topical corticosteroids. In these patients the response to topical corticosteroids was poor. CONCLUSIONS: Expression of GRbeta is increased during topical corticosteroid treatment in the lymphocytes of patients with AD and, in particular, glucocorticoid-insensitive AD is associated with increased expression of GRbeta.

0.03% Tacrolimus ointment applied once or twice daily is more efficacious than 1% hydrocortisone acetate in children with moderate to severe atopic dermatitis: results of a randomized double-blind controlled trial.

BACKGROUND: Topical corticosteroids are the usual treatment for atopic dermatitis (AD) in children but can have side-effects. OBJECTIVES: This study compared the efficacy and safety of 0.03% tacrolimus ointment applied once or twice daily over a 3-week period with the twice daily application of 1% hydrocortisone acetate (HA) ointment in children with moderate to severe AD. PATIENTS AND METHODS: Patients applied ointment daily to all affected body surface areas. The primary study endpoint was the percentage change in the modified Eczema Area and Severity Index (mEASI) between baseline and treatment end. RESULTS: Six hundred and twenty-four patients, aged 2-15 years, applied 0.03% tacrolimus ointment once daily (n = 207), twice daily (n = 210) or 1% HA twice daily (n = 207). By the end of treatment, application of 0.03% tacrolimus ointment both once or twice daily resulted in significantly greater median percentage decreases in mEASI (66.7% and 76.7%, respectively) compared with 1% HA (47.6%; P < 0.001). Furthermore, the median percentage decrease in mEASI was significantly greater for patients applying 0.03% tacrolimus twice daily compared with once daily (P = 0.007). Patients with severe AD benefited especially from twice daily application of 0.03% tacrolimus ointment compared with once daily application (P = 0.001). Transient mild to moderate skin burning occurred significantly more often in the 0.03% tacrolimus groups (P = 0.028) but resolved in most cases within 3-4 days. Laboratory parameters showed no clinically relevant changes. CONCLUSIONS: 0.03% tacrolimus ointment applied once or twice daily is significantly more efficacious than 1% HA in treating moderate-severe AD in children. Twice daily application of 0.03% tacrolimus ointment results in the greatest improvement in mEASI, and is especially effective in patients with severe baseline disease.

Demonstration of increased levels of type I collagen mRNA using quantitative polymerase chain reaction in fibrotic and granulomatous skin diseases.

Collagen changes occur in localized scleroderma, scleredema and sarcoidosis. Previous biochemical, immunohistochemical and in situ hybridization studies have revealed increased collagen synthesis in these diseases. In the present study, we measured by pro alpha 1 (I) collagen and beta-actin mRNA levels in skin punch biopsy specimens from lesional and healthy skin using a quantitative polymerase chain reaction (PCR). In this method, the targeted mRNA and a synthetic RNA as a internal standard are co-amplified together with the same primers. The amount of pro alpha 1 (I) collagen mRNA in cutaneous sarcoidosis lesions was found to be increased about two- to threefold compared with the values obtained for the healthy skin of the same two patients. In lesional skin of three patients with localized scleroderma the number of pro alpha 1 (I) collagen molecules was increased about two-fold. The beta-actin mRNA values were at the same level in the affected and unaffected skin of all the patients studied. In conclusion, a marked increase in type I collagen gene expression was seen in localized scleroderma and scleredema, leading to fibrosis of the skin, and in a granulomatous skin disease, cutaneous sarcoidosis.

Isotretinoin, tetracycline and circulating hormones in acne.

Isotretinoin, used to treat severe acne, has been shown to induce hormonal changes, especially to reduce 5 alpha-reductase in the production of the tissue-derived dihydrotestosterone (DHT) metabolite 3 alpha-Adiol G. However, the effects of isotretinoin on other pituitary, adrenal or gonadal hormones have not been thoroughly elucidated. In the present study, isotretinoin administered at a dose of 0.5 mg/kg/day for 4 weeks caused no marked changes in the serum levels of pituitary, adrenal or gonadal hormones or 3 alpha-Adiol G in patients with severe papulopustulotic acne (n = 19). After 12 weeks of therapy, there was a decrease in the levels of the precursor androgens androstenedione, testosterone and 3 alpha-Adiol G in 6/9 patients. Acne improved after 4.5 months in all but 2 male patients, who had very low serum hormone binding globulins (SHBG) and a high free androgen index (FAI). Isotretinoin did not affect the elevated LH/FSH ratio in a patient with the polycystic ovarian syndrome (PCOS); nor did it change the high FAI or low SHBG in the male patients. For comparison, tetracycline had no effects on the serum hormonal levels of patients with mild acne (n = 19) after 7 days of treatment. This study confirms that the effects of isotretinoin on the serum hormone levels are small and unlikely to be of relevance for the resolution of acne or the suppression of sebum excretion.

A homozygous nonsense mutation and a combination of two mutations of the Wilson disease gene in patients with different lysyl oxidase activities in cultured fibroblasts.

Wilson disease is a rare autosomal recessive disease of copper metabolism. The gene for Wilson disease was characterized recently and has been predicted to encode a copper-transporting ATPase highly homologous to the protein encoded by the gene of Menkes disease. In this study, the genetic mutations of two Finnish patients with Wilson disease were investigated. One patient was homozygous for a novel nonsense mutation in exon 4, while the other was a compound heterozygote. Lysyl oxidase (EC 1.4.3.13) is an extracellular copper enzyme with deficient activity in Menkes disease. The levels of lysyl oxidase activity in cultured skin fibroblasts from these Wilson disease patients were also measured.

Quantification of Pro alpha 1(I) collagen mRNA in skin biopsy specimens: levels of transcription in normal skin and in granuloma annulare.

The synthesis of type I collagen, the major component of the skin, is known to be modulated in aging and in various skin diseases and treatments. In vivo analysis of type I collagen expression, however, is difficult because of the low cell density of the dermis and the small amount of RNA obtainable from skin biopsy specimens. We present here a quantitative polymerase chain reaction method for the quantification of pro alpha 1(I) collagen mRNA in skin punch biopsy specimens. The targeted mRNA and a synthetic RNA as an internal standard were co-amplified together with the same primers. Collagen synthesis was found to decline after birth, so that the amount of pro alpha 1(I) collagen mRNA in the skin of 5- to 58-y-old donors was 17-37% of that in fetal skin. Slot-blot hybridization also indicated that the amount of pro alpha 1(I) collagen mRNA was much lower in adult skin than in fetal skin. In samples from lesional skin of two granuloma annulare patients, the number of pro alpha 1(I) mRNA molecules was increased 4- or 5-fold compared with values from nonlesional skin of the same patients. The method presented is a highly sensitive polymerase chain reaction application, requiring only very small amounts of total RNA.

Increased collagen propeptides in the skin of a scleredema patient but no change in re-epithelialisation rate.

Scleredema is a rare disease, affecting the skin connective tissue with increased amounts of collagen and glycosaminoglycans. In the present study, the collagen synthesis and re-epithelialisation rate were measured from a 64-year-old male patient, who rapidly developed extensive tightening of the skin, without any underlying disease. The skin was thickened at several sites when measured with ultrasound, and the histology revealed accumulation of glycosaminoglycans and collagen bundles. The collagen synthesis rate was measured from suction blisters induced on two different sites of the skin before the treatment and three times later up to 6 months after the treatment with a systemic steroid was started. The aminoterminal propeptide of type I collagen (PINP) was increased manifold in the affected skin when compared with the controls, indicating active collagen deposition in vivo. Systemic steroid medication with high doses (over 20 mg/d) decreased both the type I and the type III collagen propeptide levels. The time schedule of the decreases in the propeptides in the thickened, affected skin and in the clinically normal-looking skin varied, and especially in the thickened skin in the abdomen the decrease in PINP was noted only after 3 months of prednisolone therapy. When the prednisolone dose was only 10 mg daily, the propeptides were again up-regulated, perhaps reflecting the natural course of the disease. The re-epithelialisation rates at two different sites of the patient were similar to those in the controls, suggesting that even massive fibrosis with active deposition of collagen does not alter the basal rate of re-epithelialisation in the skin. In conclusion, collagen synthesis is markedly elevated in scleredema, leading to fibrosis of the skin. A recently developed method utilizing assays of collagen propeptides from suction blister fluid allows monitoring of the collagen synthesis and detection of changes in the collagen synthesis during the treatment of fibrotic disorders.

Systemic oral isotretinoin therapy and flow rate, pH, and matrix metalloproteinase-9 activity of stimulated saliva.

Systemic retinoids are known to cause dryness of the mouth and changes in oral and lip mucosa. The purpose of this study was to evaluate changes in salivary variables during treatment with oral isotretinoin in patients receiving the drug for 3 months for cutaneous acne. Patients were examined 1 month after initiation of medication and approximately 3.7 months after its discontinuation. Salivary flow and pH could be measured in 8 and the relative amount of matrix metalloproteinase-9 (MMP-9) of stimulated saliva in 17 patients. The mean flow rate of stimulated saliva was lower during medication than at control examination (P = 0.0277), but no change in the mean pH value was observed during medication. The mean activity of MMP-9 during medication was higher than at control examination (P = 0.0442). The enzyme activity increased in 13 of 17 and decreased in 4 of 17 cases.

Synthesis and degradation of connective tissue macromolecules in pachydermoperiostosis (PDP): evidence for altered processing of plasminogen activator inhibitor-1 (PAI-1).

Pachydermoperiostosis (PDP) is a hereditary disease with hyperostosis, clubbing of fingers, coarse skin and thickening of bones. Previous studies have disclosed some abnormality in the connective tissue in these patients. The purpose of the present study was to investigate connective tissue pathology in one family with PDP using fibroblast cultures. Fibroblastic cells were established from both the affected and healthy looking skin of 2 patients with PDP, and the expression of types I and III collagen, 92 kDa and 72 kDa gelatinases, metalloproteinase inhibitor (TIMP-1), human retinoic acid receptor and transforming growth factor beta (TGF beta) was analyzed. The modulation of glycoprotein synthesis, and of plasminogen activators and their inhibitors by TGF beta in vitro were also studied. The results indicated that collagen genes and gelatinases were similarly expressed in PDP and control cells, as well as the human retinoic acid receptor. TGF beta stimulated, both in PDP cells and normal cells, the synthesis of fibronectin, procollagen and plasminogen activator inhibitor-l (PAI-1), but qualitative differences could not be found. Proteolytically processed forms of PAI-1 were detected in PDP cell lines.

Pachydermoperiostosis: analysis of the connective tissue abnormality in one family.

BACKGROUND: Pachydermoperiostosis (PDP) is a rare hereditary disease characterized by hyperostosis, clubbing of fingers, coarse skin, and abnormalities in other organs, such as the gastrointestinal tract. Previous studies have disclosed several abnormalities in the connective tissue in these patients. OBJECTIVE: The purpose of the study was to investigate connective tissue abnormalities in one family with PDP. METHODS: Clinical features were evaluated; x-ray, immunohistochemical, and electronmicroscopic studies were performed; and markers of collagen metabolism and lysosomal enzymes were determined. RESULTS: Immunohistochemical and ultrastructural studies revealed accumulation of tenascin, glycosaminoglycans, and fibrillar material in apparently disorganized microfibrils of elastic fibers. Osteocalcin levels in the serum were increased, but synthesis and degradation markers of collagen in the serum were not altered. No evidence of a lysosomal enzyme deficiency was found. CONCLUSION: Acidic mucopolysaccharides and some fibrillar material accumulate in the dermis of patients with PDP. Increased levels of osteocalcin in serum indicate higher osteoblastic activity. Markers of synthesis and degradation of collagen were not altered.

Effects of systemic isotretinoin on serum markers of collagen synthesis and degradation.

In the present investigation, collagen synthesis and degradation were studied by measuring the carboxyterminal propeptide of type I procollagen (PICP), the aminoterminal propeptide of type III procollagen (PIIINP) and a type I collagen-specific degradation peptide (ICTP) in the sera of 43 male patients, treated for acne with isotretinoin or with tetracycline. The values were compared with those observed in 24 acne patients without treatment and in healthy controls. The treatment with isotretinoin did not seem to affect these parameters in a cross-sectional setting, whereas tetracycline treatment was associated with slightly decreased levels of ICTP. Since there were marked variations in the PICP, PIIINP and ICTP levels between individual subjects, a follow-up study, including male and female patients, others than in the first part of the study, was conducted. Two other biochemical markers of bone metabolism, osteocalcin, reflecting osteoblastic activity, and tartrate-resistant acid phosphatase (TRAP), reflecting osteoclastic activity, were also analyzed. In females, all these parameters were lower than in males. In addition, the changes in females were more pronounced; in particular, PIIINP and TRAP were significantly increased in females during retinoid treatment (p < 0.05 and p < 0.01, respectively). Importantly, no increase was found in the synthesis of type I collagen during retinoid treatment, suggesting that the commonly used retinoid dosages do not stimulate the synthesis of type I collagen in vivo.

Comparison of muscle-derived serum carbonic anhydrase III and myoglobin in dermatological patients: effects of isotretinoin treatment.

The serum levels of muscle-specific serum carbonic anhydrase III (S-CAIII) and myoglobin (S-Myo) were analyzed in various male dermatological patients of the same age. The mean levels of S-CAIII and S-Myo were essentially similar in patients with acne, psoriasis vulgaris, atopic eczema and tinea, suggesting that common dermatological diseases do not affect the serum levels of the muscle markers. Increased levels of S-CAIII, which is specific for skeletal muscle cells, were found in the acne patients who had been treated with isotretinoin. However, when S-CAIII and S-Myo were studied in 24 patients (16 males, 8 females) before and during isotretinoin treatment, no constant increases in these markers could be observed. When individual patients were followed for several months, transient increases or decreases could be observed. The changes in S-CAIII, or S-Myo, did not correlate with the dose of isotretinoin, nor with the duration of the treatment. The results suggest that systemic isotretinoin does not specifically affect skeletal or myocardial muscles. The increases in these markers observed in the course of dermatological diseases and isotretinoin treatment are obviously due to other factors, such as exercise.

Scleredema and paraproteinemia. Enhanced collagen production and elevated type I procollagen messenger RNA level in fibroblasts grown from cultures from the fibrotic skin of a patient.

An edematous rash developed on the abdominal skin of a 76-year-old woman who had had diabetes mellitus for ten years. Some months later, the affected skin became thickened and indurated. Histopathologic examination revealed marked dermal fibrosis with excessive deposition of collagen. The patient also had IgA (k-type) paraproteinemia. Fibroblast cultures from the affected and unaffected skin were studied for collagen metabolism. Procollagen synthesis was elevated about sixfold on fibroblasts derived from the affected skin. A similar increase was detected in messenger RNA (mRNA) levels using a complementary DNA clone specific for human pro alpha 1(l) collagen mRNA. The elevated mRNA level could be the result of increased transcriptional activity of collagen genes or decreased degradation of collagen mRNAs. Our findings suggest that increased collagen deposition may account for the marked dermal fibrosis that we observed in this patient.

Paraproteinemia in patients with scleredema. Clinical findings and serum effects on skin fibroblasts in vitro.

Four patients with paraproteinemia and scleredema were studied. Histologic features included marked thickening and fibrosis of the dermis and subcutis. Variable amounts of mucin deposits were detected in the interfibrillar spaces. Serum from one patient significantly stimulated collagen production in normal skin fibroblast cultures, whereas serum from another patient stimulated collagen production in autologous cell cultures. Moreover, serum from one patient stimulated the [35S]sulfate incorporation into the fibroblasts. Circulating serum factors, possibly related to the paraprotein, may enhance the synthesis of extracellular macromolecules by dermal fibroblasts in these patients, thus providing a mechanism for dermal fibrosis.

Pituitary function and DHEA-S in male acne and DHEA-S, prolactin and cortisol before and after oral contraceptive treatment in female acne.

Pituitary function (TRH-LHRH stimulation test) was investigated in male acne patients and serum levels of dehydroepiandrosterone sulphate (DHEA-S), sex hormone binding globulin (SHBG) and other biochemical parameters were investigated in male acne patients and in female acne patients before and after treatment with an oral contraceptive. The TRH-LHRH stimulation test was performed with 15 male patients suffering from severe cystic acne and 7 healthy volunteers. Basal and stimulated prolactin, LH and FSH levels were statistically similar in the patients and control groups. However, the stimulated LH levels of the patients were 60% higher than those in controls. SHBG levels were significantly) higher in the patient group compared to those in the control group. Thirty-three female acne patients were randomly divided into two groups and treated for six months with an oral contraceptive containing 0.030 mg ethinylestradiol (EE) plus 0.150 mg levonorgestrel or 0.150 mg levonorgestrel. After six months' treatment a 30% decrease in DHEA-S levels were observed in the desogestrel/EE group and a 15% decrease in the levonorgestrel/EE group; the difference was not statistically significant. At the same time serum total cortisol increased by 75-100% and free testosterone fell by 30-40% in both groups, whereas SHBG elevated 250% in the desogestrel/EE group and 30% in the levonorgestrel/EE group. Acne improved significantly in both groups, desogestrel/EE showing greater improvement. A decrease in SHBG and increase in DHEA-S levels appear to be the most common hormonal changes in acne. Oral contraceptive treatment induces an increase in SHBG and decrease in DHEA-S and also improves acne.

PIP: This study investigated pituitary function in male acne patients and serum levels of dehydroepiandrosterone sulphate (DHEA-S), sex hormone binding globulin (SHBG), and other biochemical parameters in male acne patients and in female acne patients before and after oral contraceptive (OC) treatment. Among males, basal and stimulated prolactin, luteinizing hormone (LH), and follicle-stimulating hormone (FSH) levels were similar among acne cases and controls; however, stimulated LH levels were 60% higher among cases and SHBG levels were 60% higher among cases and SHBG levels were statistically higher. The 33 female acne patients were treated for 6 months with an OC containing 0.03 mg ethinyl estradiol plus 0.15 mg levonorgestrel or 0.15 desogestrel. After 6 months, a 15% and a 30% decrease, respectively, was observed in DHEA-S levels in the 2 groups. Serum total cortisol increased by 75-100% and free testosterone fell by 30-40% in both groups, while SHBG elevated 250% in the desogestrel/ethinyl estradiol group and 30% in the levonorgestrel/ethinyl estradiol group. Although acne improved in both groups, the improvement was greatest among cases receiving a desogestrel-containing OC. These findings suggest that low SHBG and elevated DHEA-S are the hormonic alterations most often seen in cases of severe acne, although pretreatment DHEA-S levels did not correlate with the severity of the acne. It is concluded that OCs improve acne by increasing SHBG and decreasing DHEA-S levels.

Anetoderma: biochemical and ultrastructural demonstration of an elastin defect in the skin of three patients.

Three patients with localized cutaneous lesions characteristic of anetoderma were studied. Clinically, the onset of the disease was between the ages of 17 and 25, and numerous flaccid, saclike skin lesions developed over several subsequent years. Histologically, the lesions were characterized by paucity and fragmentation of the elastic fibers. Electron microscopy demonstrated that the elastic fibers, both in papillary and deep reticular dermis in the lesional skin, were fragmented and irregular in appearance. The concentration of elastin, determined by a radioimmunoassay of desmosine, an elastin-specific cross-link compound, was markedly reduced in the lesions, as compared with unaffected skin from the same patients or with normal skin from unrelated control subjects. In contrast, the concentrations of hydroxyproline, an index of collagen, or deoxyribonucleic acid (DNA), a measure of cellularity, were not changed in the lesions. Thus, the results indicate that in the three patients studied, the elastic fibers are defective and reduced in quantity. These observations suggest that the deficiency of elastin in the dermis may lead to development of the cutaneous lesions of anetoderma.

Serum total and unbound testosterone and sex hormone binding globulin (SHBG) in female acne patients treated with two different oral contraceptives.

Serum total an unbound testosterone (T) and sex hormone binding globulin (SHBG) levels were studied in fifty-four female acne patients before treatment and during the treatment by two different oral contraceptives, the other containing 0.150 mg desogestrel plus 0.03 mg EE and the other 0.150 mg levonorgestrel plus 0.03 mg EE. Pretreatment values were abnormal in 57% of the patients. A borderline significant correlation between the severity of acne and SHBG was found. Ater six months' treatment a 250% increase in SHBG was seen in desogestrel/EE group and no significant change in SHBG in levonorgestrel/EE group. However, at the same time serum free testosterone fell 60% in both treatment groups. SHBG cannot be the only regulator of serum free testosterone. Acne improved significantly in both treatment groups. It is likely that the improvement was in connection with the free testosterone decrease and the improvement was better in the desogestrel/EE group where also SHBG elevation was seen.

PIP: Serum total and unbound testosterone (T) and sex hormone binding globulin (SHBG) levels were studied in 54 female acne patients before and during the treatment by 2 different oral contraceptives, 1 containing 0.150 mg desogestrel + 0.03 mg ethinyl estradiol (EE) and the other 0.150 mg levonorgestrel + 0.03 mg EE. Pretreatment values were abnormal in 57% of the patients. A borderline significant correlation between the severity of acne and SHBG was found. After 6 months of treatment, a 250% increase in SHBG was seen in the desogestrel/EE group and no significant change in SHBG level in the levonorgestrel/EE group. However, at the time same, serum free testosterone fell 60% in both treatment groups. SHBG cannot be the only regulator of serum free testosterone. Acne improved significantly in both treatment groups. It is likely that the improvement was the result of the free testosterone decrease and the imnprovement was greater in the desogsertel/EE group where SHBG elevation was also seen.

Types I and III collagens and the activities of prolyl hydroxylase and galactosylhydroxylysyl glucosyltransferase in skin lesions of tuberous sclerosis.

Collagen synthesis and the ratios of collagen types I and III were assayed from the skin lesions of five subjects with tuberous sclerosis. Collagen synthesis, measured by the activities of prolyl hydroxylase and galactosylhydroxylysyl glucosyltransferase, was clearly increased in three angiofibromas of these patients and in one soft tumour of the face, but it was unchanged in shagreen patches. The total collagen content was decreased in angiofibromas, indicating either increased turnover of collagen or an increased amount of cellular or other macromolecular elements in these lesions. The proportions of types I and III collagens, estimated by cyanogenbromide digestion and SDS-gel electrophoresis, were 80-90% and 10-20%, respectively, in all samples except two angiofibromas, in which the relative amount of type III collagen was increased. This may indicate that angiofibromas of tuberous sclerosis are heterogenous with respect to the collagen types they contain, and that there may be disturbed cell growth or collagen synthesis, with individual variation from case to case.