Muscular response to the first three months of deflazacort treatment in boys with Duchenne muscular dystrophy.

OBJECTIVE: Duchenne muscular dystrophy (DMD) patients are often treated with glucocorticoids; yet their precise molecular action remains unknown. METHODS: We investigated muscle biopsies from nine boys with DMD (aged: 7,6±2,8 yrs.) collected before and after three months of deflazacort treatment and compared them to eight healthy boys (aged: 5,3±2,4 yrs.). mRNA transcripts involved in activation of satellite cells, myogenesis, regeneration, adipogenesis, muscle growth and tissue inflammation were assessed. Serum creatine kinase (CK) levels and muscle protein expression by immunohistochemistry of selected targets were also analysed. RESULTS: Transcript levels for ADIPOQ, CD68, CDH15, FGF2, IGF1R, MYF5, MYF6, MYH8, MYOD, PAX7, and TNFα were significantly different in untreated patients vs. normal muscle (p⟨0.05). Linear tests for trend indicated that the expression levels of treated patients were approaching normal values (p⟨0.05) following treatment (towards an increase; CDH15, C-MET, DLK1, FGF2, IGF1R, MYF5, MYF6, MYOD, PAX7; towards a decrease: CD68, MYH8, TNFα). Treatment reduced CK levels (p⟨0.05), but we observed no effect on muscle protein expression. CONCLUSIONS: This study provides insight into the molecular actions of glucocorticoids in DMD at the mRNA level, and we show that multiple regulatory pathways are influenced. This information can be important in the development of new treatments.

Interferon-alfa treatment of facial infantile haemangiomas: with emphasis on the sight-threatening varieties. A clinical series.

PURPOSE: To report on the efficacy of systemic Interferon-alfa therapy in controlling the endothelial proliferation in expanding facial haemangiomas of infancy threatening the eye. METHODS: The series comprised 9 subjects with facial haemangiomas of rapid growth within the first few months of life. All haemangiomas had relation to an eyelid and affected the visual access to the eye. At least three tumours were also intraorbital, and in one case the glottis was affected. Interferon-alfa was given subcutaneously in daily doses of 3 mio units/m2. Ophthalmic follow-up including ultrasound evaluation was done, initially with intervals of 4-8 weeks. RESULTS: Interferon-alfa was given over 9-41 weeks (median duration 22 weeks) in cumulated doses of 63-287 million units. Progression was arrested in all and with a subsequent tumour regression considered accelerated as judged from earlier clinical giant haemangioma experience. A control series was not possible. One patient had systemic prednisone added over some weeks. Two infants reacted with a flu-like malaise. A premature infant developed a slight spastic diplegia. CONCLUSIONS: In the 9 infants with growing ophthalmic haemangiomas we recorded a good response to systemic Interferon-alfa administered as daily doses by subcutaneous injection. In general the treatment appeared well tolerated. During clinical follow-up diagnostic ultrasound evaluation (the depth dimension) proved helpful; in particular there was a sparing effect on CT and MRI scans which require general anaesthesia.

[Diagnosis of pyruvate kinase deficiency]. / Diagnostik af pyruvatkinasemangel.

Two children with nonspherocytic anaemia and pyruvate kinase deficiency had their diagnosis delayed due to normal initial estimations of enzyme activity. Repeated analyses involving other glycolytic enzymes documented an increased glucose-6-phosphate dehydrogenase activity as a function of high reticulocyte counts. The pyruvate kinase activity showed subnormal values. By simple comparison of both enzyme activities a true pyruvate kinase activity of below 50% was estimated thus rendering the diagnosis of pyruvate kinase deficiency highly probable. Analyses of more than one glycolytic enzyme should be performed in young children with otherwise unexplained haemolysis and associated high reticulocyte counts.

Ulcerated haemangioma successfully treated with interferon alfa-2b and topical granulocyte-macrophage colony-stimulating factor.

OBJECTIVE: We tested whether healing and regression could be promoted by granulocyte-macrophage colony-stimulating factor and interferon in a 6-month-old girl with an ulcerated haemangioma resistant to systemic steroid therapy. METHODS: Interferon alfa-2b (Introna, Schering-Plough) was given subcutaneously once daily at a dose of 3 million units/m2 for 5.5 months and rhGM-CSF (Molgramostim, Leucomax; Sandoz/Schering-Plough) 3.33 units was applied on the wound surfaces once and concomitantly with the initiation of interferon therapy. RESULTS: The ulcer healed completely within 1 month and the haemangioma almost totally regressed within 6 months. CONCLUSION: Local application of rhGM-CSF appears to be effective in promoting ulcer healing in an ulcerated haemangioma otherwise responding to interferon treatment.

Human red cell Aquaporin CHIP. II. Expression during normal fetal development and in a novel form of congenital dyserythropoietic anemia.

Channel-forming integral protein (CHIP) is the archetypal member of the Aquaporin family of water channels. Delayed CHIP expression was shown recently in perinatal rat (Smith, B. L., R. Baumgarten, S. Nielsen, D. Raben, M. L. Zeidel, and P. Agre. 1993. J. Clin. Invest. 92:2035-2041); here we delineate the human patterns. Compared with adult, second and third trimester human fetal red cells had lower CHIP/spectrin ratios (0.72 +/- 0.12, 0.94 +/- 0.22 vs 1.18 +/- 0.11) and reduced osmotic water permeability (0.029, 0.026 vs 0.037 cm/s); CHIP was already present in human renal tubules by the second trimester. A patient with a novel form of congenital dyserythropoietic anemia (CDA) with persistent embryonic and fetal globins and absent red cell CD44 protein was studied because of reduced CHIP-associated Colton antigens. Novel CDA red cells contained < 10% of the normal level of CHIP and had remarkably low osmotic water permeability (< 0.01 cm/s), but no mutation was identified in Aquaporin-1, the gene encoding CHIP. These studies demonstrate: (a) unlike rat, human CHIP expression occurs early in fetal development; (b) red cell water channels are greatly reduced in a rare phenotype; and (c) disrupted expression of red cell CHIP and CD44 suggests an approach to the molecular defect in a novel form of CDA.

A novel form of congenital dyserythropoietic anemia associated with deficiency of erythroid CD44 and a unique blood group phenotype [In(a-b-), Co(a-b-)].

We have used a panel of well-characterized monoclonal antibodies (MoAbs) to examine the blood cells of a patient with a novel form of congenital dyserythropoietic anemia (CDA) characterized by intra-erythroblastic and intra-erythrocytic membranous inclusions. Twelve antibodies defining three nonoverlapping epitope groups on the extracellular domain of CD44 all failed to react with the red blood cells (RBCs) of the patient. A rabbit antibody to the cytoplasmic domain of CD44 from normal RBCs failed to react with the patient's RBC ghosts. In contrast, the patient's lymphocytes, granulocytes, and monocytes showed apparently normal CD44 expression. Bone marrow preparations stained with CD44 antibodies and visualized with 125I antimouse Ig (F(ab')2) followed by autoradiography showed positive staining of lymphocytes and myeloid cells but not of most orthotolidine-positive erythroblasts. The patient's RBCs also gave weaker than normal reactions with MoAbs of anti-LWab specificity while MoAbs to glycophorins A, B, and C, Rh polypeptides, CD47, CD55, CD58, CD59, acetylcholinesterase, and Lutheran and Kell glycoproteins all gave normal reactions. Agglutination tests with human blood grouping sera demonstrated that the RBCs of the patient have the unique phenotype In(a-b-), Co(a-b-) and that they also lack the high incidence RBC antigen AnWj. The phenotype In(a-b-) would be expected because these antigens are known to be expressed on CD44. There is also some evidence associating the AnWj antigen with CD44. However, the CO blood group locus is on chromosome 7p whereas that for CD44 is on chromosome 11p. Quantitative binding assays using 125I-labeled Fab fragments of CD44 antibodies did not show any evidence for reduced levels of CD44 on RBCs from the parents of the patient or from her unaffected sister. The parents and sister had the common Colton blood group phenotype [Co(a+b-)]. Neither deficiency of CD44 nor absence of Colton antigens are general features of CDA because erythrocytes from patients with CDA I, CDA II, CDA III, and two other unclassified CDAs had normal expression of CD44 and normal Colton blood group phenotypes. Further analysis of the defect(s) present in the patient's erythroid cells may provide useful information regarding membrane assembly and the regulation of differentiation in normal erythroid cells.

The role of Mycobacteria Other Than Tuberculosis (MOTT) in patients with cystic fibrosis.

The purpose of this study was to estimate the frequency of and evaluate the clinical impact of pulmonary mycobacterial infections among cystic fibrosis (CF) patients. 185 CF patients aged 2.2-38.5 years were screened by sputum samples and by intracutaneous skin tests against tuberculin and sensitins produced from Mycobacterium chelonae subsp. abscessus, M. avium, M. intracellulare and M. scrofulaceum (the MAIS complex). The skin tests towards the sensitins in BCG-vaccinated patients (n = 60) were significantly influenced by the vaccination. 26 of the remaining 125 non-vaccinated patients had > or = 1 positive skin test (95% confidence limits 15-29%). The majority reacted against the MAIS complex. However, the reactions were similar to those of healthy siblings and an age-matched control group. Moreover, the lung function, growth and HbA1c were similar among skin test positive and negative patients. Three patients had repeated positive sputum cultures, the point prevalence being 1.6% (M. intracellulare, n = 2 and M. chelonae subsp. abscessus, n = 1). During the subsequent 4 years, 4 additional patients with M. chelonae subsp. abscessus were identified. Based on clinical observations, 5 of the infected patients were considered asymptomatic, while 2 might have been symptomatic. In 1 patient, M. chelonae subsp. abscessus disappeared spontaneously. Despite intensive treatment with new antibiotics against Mycobacteria Other Than Tuberculosis (MOTT) in 4 patients, the mycobacteria were not eradicated. In conclusion, MOTT infection was rare and the clinical impact difficult to prove. Treatment should focus on clinical improvement in the individual patient suspected of suffering from significant symptomatic infection. Eradication of the bacteria should not be expected.

Expression of embryonic zeta-globin and epsilon-globin chains in a 10-year-old girl with congenital anemia.

A 10-year-old Danish girl with congenital anemia is described. At birth, she had severe anemia and erythroblastosis and was transfused a number of times during the first year. The need for transfusions has since declined steadily. Her reticulocyte counts varied between 2% and 15%, and her bone marrow aspirate showed some dyserythropoietic features. Her hemoglobin F level was consistently elevated, up to as much as 41%. Her erythrocytes had a normal level of I antigen but an undetectable level of i antigen. Moreover, embryonic zeta-globin and epsilon-globin chains were present in some of her circulating erythrocytes. These findings may represent the manifestations of a new variant of congenital anemia.

[Physiopathologic mechanisms behind eye symptoms in primary tumors of the pineal body]. / Patofysiologiske mekanismer bag øjensymptomer ved primaere tumorer i corpus pineale.

Primary tumors of the pineal body can produce dyscoordinative movements of the eye, pupillary dilatation, paralysis of adduction during convergence and nystagmus. Obstruction of the aqueduct can cause hydrocephalus, increased intracranial pressure and papilledema. Diabetes insipidus may be a presenting symptom. Pinealocytes and the photoreceptors of the eye contain several autoantigens. In man, the best known is the S-antigen. This antigen can be detected in the cerebrospinal fluid of patients with primary tumors of the pineal body. The S-antigen, and possibly other related autoantigens, can elicit an autoimmune mediated reaction causing inflammatory eye symptoms. This recently described paraneoplastic neurologic syndrome shares properties in common with other known cancer-associated ophthalmologic syndromes characterised by rapid development of eye symptoms, rapid loss of sight and by eye manifestations prior to evident appearance of symptoms related to primary tumor growth. A primary tumor of the pineal body should be considered in patients where a monosymptomatic uveoretinitis presents without associated provoking factors. Furthermore, analyses of S-antigen in the spinal fluid can be useful in the clinical diagnosis of the same primary tumors.

[Treatment of hemangioma in children with recombinant interferon]. / Behandling af haemangiom hos børn med rekombinant interferon.

Pulmonary haemangiomatosis and large haemangiomas endangering vital structures and with a potential association of consumptive coagulopathy are rare but severe diseases in infancy and childhood. Therapy with glucocorticoids, cytostatics and surgical intervention has not proved satisfactory. From experience with Kaposi's sarcoma, about 20 children with severe forms of haemangiomatous disease have now been treated with alpha-interferon with promising results. Mechanisms of action are unknown, but these could include a cytostatic effect, inhibition of the abnormal proliferation of endothelial cells, fibroblasts and smooth muscle cells. Furthermore, alpha-interferon could inhibit the paracrine and autocrine effects of locally produced growth factors. Also, alpha-interferon could promote the production of prostacyclin, which inhibits platelet aggregation, leading to reduced damage and consumption of platelets. Long term treatment is mandatory. Initial daily doses of 2 to 5 million U/m2 are administered subcutaneously. Side effects in this dosage interval are uncommon in childhood. Future studies are needed to establish exact guidelines for the use of alpha-interferon in childhood, but the clinical experience hitherto obtained points to the fact that treatment is effective in severe forms of haemangiomas and pulmonary haemangiomatosis.

Concurrent uveoretinitis and pineocytoma in a child suggests a causal relationship.

Uveoretinitis was observed in a 9-year-old girl 6 months prior to the clinical appearance of a pineal tumour. Surgical removal was not successful but biopsy revealed a parenchymal neoplasm with differentiated pinealocytes and absent mitotic activity. Some of the tumour cells contained S-antigen, rhodopsin, and serotonin. Systemic glucocorticoid therapy followed by radiation therapy caused considerable reduction in size of the tumour and a complete normalisation of all eye symptoms. This report demonstrates for the first time that a pineocytoma can occur together with uveoretinitis in humans. The latter resembles the experimentally induced autoimmune uveoretinitis described in animals. It is speculated that the retinitis might reflect an autoimmune response to S-antigen present in some tumour cells of the pineocytoma.

Recurrent nodular haemangiomas in Klippel-Trénaunay syndrome.

A one-year-old child had hypertrophy of the left leg and an unusual constellation of a naevus flammeus and superficial enlarged veins of the trunk together with successive appearance and involution since birth of numerous nodular elements located in the naevus and in the surrounding normal skin. Microscopic examination of these elements showed haemangiomas with capillaries, cavernous channels and lymphangiomatous components. The benign nature of transient nodular elements located to the trunk and the lack of associated visceral vascular malformations in the Klippel-Trénaunay syndrome are documented.

[Neurocysticercosis. Severe neurocysticercosis in a child]. / Neurocysticerkose. Svoer neurocysticerkose hos et barn.

A case of neurocysticercosis with progressive severe neurological symptoms is described. The patient was a Turkish girl aged 4 1/2 years who had experienced intermittent neurological symptoms for two years. Rapid diagnosis and treatment with praziquantel and corticosteroid resulted in complete restitution.

Primary hyperoxaluria type 1: clinical manifestations in infancy and prenatal diagnosis.

A 9-month-old Pakistani boy of consanguineous parents presented with uraemia preceded by pyuria from 5 weeks of age. He had no history of renal calculi or macroscopic haematuria. Renal biopsy revealed severe calcium oxalate deposition in the tubuli and fibrosis of the interstitial tissue. Liver biopsy demonstrated complete absence of alanine: glyoxylate aminotransferase catalytic activity and immunoreactive protein compatible with a diagnosis of primary hyperoxaluria type 1. He died at the age of 11 months, just before liver transplantation was made possible. Fetal liver biopsy in the mother's subsequent pregnancy showed normal enzymatic activity. Early detection and early replacement of the missing enzyme by liver transplantation are considered to be crucial for the survival of severely affected infants with the acute neonatal form of primary hyperoxaluria type 1. Persistent pyuria could be an early sign of renal damage secondary to accumulation of oxalate crystals.

Phenotypical and functional characterization of double-negative (CD4-CD8-) alpha beta T-cell receptor positive cells from an immunodeficient patient.

We have characterized CD4-CD8- double-negative (DN) alpha beta TCR+ T cells from a patient with immunodeficiency, lymphocytosis, lymphadenopathy, and hepatosplenomegaly. The majority of peripheral blood lymphocytes were DN alpha beta TCR+ T cells as evaluated by FACS and biochemical analysis. The DN T cells showed the following phenotype: alpha beta TCR+, gamma delta TCR-, CD2+, CD3+, CD4-, CD5+, CD7-, CD8-, CD16-, CD25-, CD26-, CD28+, CD45RO-, CD45RA+, CD57+, and HLA-DR+. Both southern blot analysis of TCR genes and FACS analysis applying a panel of V beta and V alpha monoclonal antibodies (MoAbs) indicated a polyclonal T-cell expansion. Thymic biopsy showed normal histology, whereas lymph node biopsy samples showed altered histological and immunohistological patterns with markedly expanded paracortical areas containing the DN T cells of the same phenotype as found in peripheral blood T cells. In functional studies, the DN T cells showed a profoundly reduced proliferative response upon stimulation with mitogens as well as MoAbs against the TCR/CD3 complex, CD2, and CD28, respectively. Addition of exogenous interleukin-2 (IL-2) only minimally augmented the proliferative response. In contrast, the addition of a combination of Ca2+ ionophore and phorbol 12-myristate 13-acetate (PMA) restored the proliferative response of the DN T cells to almost normal levels. This observation strongly suggests that the protein kinase C activity of the DN T cells was intact, but that the normal mechanism for transmembrane signal transduction was impaired in these unusual DN T cells.

Congenital dyserythropoietic anaemia with novel intra-erythroblastic and intra-erythrocytic inclusions.

A hitherto undescribed form of congenital dyserythropoietic anaemia is reported. The patient was severely anaemic and hydropic at birth and is now 8 years old. She has a moderate normochromic normocytic anaemia. HbF level of 50%, reticulocyte count of 5-12% and hyperbilirubinaemia. Bone marrow smears showed intense normoblastic erythroid hyperplasia with morphological evidence of dyserythropoiesis; the most common dysplastic features were basophilic stippling of polychromatic erythroblasts and erythrocytes and marked abnormalities of nuclear shape in polychromatic erythroblasts. Electron microscope studies showed that some polychromatic erythroblasts and several erythrocytes contained inclusions which were rounded, elongated or irregular in outline or were doughnut-shaped. These inclusions consisted of compact masses of tubules and saccules which may represent smooth endoplastic reticulum together with Golgi cisternae. The ultrastructural studies also revealed peculiar membrane-bound cylindrical structures in a rare late erythroblast, and phagocytosed erythroblasts within some macrophages. The technique of combined Feulgen microspectrophotometry and 3H-thymidine autoradiography demonstrated a pile-up of early polychromatic erythroblasts in the G1 and G2 phases of the cell cycle, indicating a prolongation of, or an arrest at, these phases. Furthermore, nearly a quarter of all erythroblasts failed to incorporate 3H-leucine into protein. Thus the anaemia appeared to be due to a combination of disordered erythroblast function, increased ineffectiveness of erythropoiesis and peripheral haemolysis. The primary defect may be an excessive synthesis or impaired degradation of intracytoplasmic membranes.

Intravenous immunoglobulin: a single-blind trial in children with Lennox-Gastaut syndrome.

The antiepileptic effect of intravenous immunoglobulin (Sandoglobulin, Sandoz) was investigated in Lennox-Gastaut syndrome by an add-on, placebo-controlled, single-blind trial. Ten patients, aged 4-14 years, with insufficient response to conventional anticonvulsive therapy received placebo and Sandoglobulin 400 mg/kg two times each with an interval of two weeks. The washout period was four weeks and the total observation period 14 weeks, during which parents daily registered number and type of seizures. EEG, in vitro lymphocyte transformation tests and concentrations of immunoglobulins including IgG subclasses were evaluated before and after active treatment. Two children showed an immediate reduction in their high-frequency and invariable seizure activity from 42% to 100% and a less abnormal EEG. In addition, general well-being and intellectual performance was improved. The strongest response was observed in one child with a concomitant finding of a low level of IgG2, the only abnormal immunologic test in this study. The remaining 8 children, who had either a high or a low but variable seizure frequency showed no immediate change as EEG and their general condition was unaffected. We conclude that intravenous immunoglobulin had an immediate and pronounced effect on break-through seizure activity and a simultaneous neurophysiologic effect in 20% of our patients with Lennox-Gastaut syndrome. The effect was not confined to patients with immunologic abnormalities.

Lethal autosomal recessive arthrogryposis multiplex congenita with whistling face and calcifications of the nervous system.

Three children from a sibship of four had congenital contractures, scarce facial expressions, central nervous system dysfunction, and early death. Extensive deposits of calcium compounds were found at postmortem examination of the nervous system and of skeletal muscle. The disorder in these sibs is presumably inherited as an autosomal recessive trait. The metabolic basis for the calcium deposition has yet to be discovered.