Mitochondrial neurogastrointestinal encephalomyopathy: Clinical and biochemical impact of allogeneic stem cell transplantation in a Greek patient with one novel TYMP mutation.

We describe the case of a Greek female patient with the Classic form of the ultra- rare and fatal autosomal recessive disorder Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) and the impact of allogeneic hematopoietic stem cell transplantation on the biochemical and clinical aspects of the disease. The patient presented at the age of 15 years with severe gastrointestinal symptoms, cachexia, peripheral neuropathy and diffuse leukoencephalopathy. The diagnosis of MNGIE disease was established by the increased levels of thymidine and deoxyuridine in plasma and the complete deficiency of thymidine phosphorylase activity. The novel c.[978dup] (p.Ala327Argfs*?) variant and the previously described variant c.[417 + 1G > A] were identified in TYMP. The donor for the allogeneic hematopoietic stem cell transplantation was her fully compatible sister, a carrier of the disease. The patient had a completely uneventful post- transplant period and satisfactory PB chimerism levels. A marked and rapid decrease in thymidine and deoxyuridine plasma levels and an increase of the thymidine phosphorylase activity to the levels measured in her donor sister was observed and is still present sixteen months post-transplant. Disease symptoms stabilized and some improvement was also observed both in her neurological and gastrointestinal symptoms. Follow up studies will be essential for determining the long term impact of allogeneic hematopoietic stem cell transplantation in our patient.

Dihydropyrimidine dehydrogenase deficiency caused by a novel genomic deletion c.505_513del of DPYD.

Dihydropyrimidine dehydrogenase (DPD) deficiency is an autosomal recessive disorder of the pyrimidine degradation pathway. In a patient presenting with convulsions, psychomotor retardation and Reye like syndrome, strongly elevated levels of uracil and thymine were detected in urine. No DPD activity could be detected in peripheral blood mononuclear cells. Analysis of the gene encoding DPD (DPYD) showed that the patient was homozygous for a novel c.505_513del (p.169_171del) mutation in exon 6 of DPYD.

Identification of two novel mutations C79X and R235Q in the dihydropyrimidine dehydrogenase gene in a patient presenting with hematuria.

A patient with hematuria was shown to have thymine-uraciluria. The dihydropyrimidine dehydrogenase (DPD) activity in peripheral blood mononuclear cells was 0.16 nmol/mg/h; controls: 9.9 +/- 2.8 nmol/mg/h. Analysis of DPYD showed that the patient was compound heterozygous for the novel mutations 237C > A (C79X) in exon 4 and 704G > A (R235Q) in exon 7. The nonsense mutation (C79X) leads to premature termination of translation and thus to a non-functional protein. Analysis of the crystal structure of pig DPD suggested that the R235Q mutation might interfere with the binding of FAD and the electron flow between the NADPH and the pyrimidine substrate site of DPD.

[Evaluation and validation of a perinatal death audit by means of feedback to the caregivers]. / Evaluatie en validatie van perinatale-sterfte-audit door terugkoppeling naar zorgverleners.

OBJECTIVE: To evaluate a perinatal audit procedure by communicating the results to the caregivers (midwives and obstetricians) involved, in order to determine whether the audit led to specific suggestions for improving practice and whether evaluation of the panel assessments by caregivers leads to a different evaluation of the audit process. DESIGN: Descriptive evaluation study. METHOD: Because of privacy regulations, the results of a recently published audit concerning perinatal mortality were reported at an aggregated level. At their own request, two participating hospitals received panel assessment reports of their own cases. The audit procedure, the 77 panel assessments and the care provided were then evaluated during closed meetings with the caregivers affiliated to the respective hospitals. RESULTS: In two audited cases of mortality the caregivers judged the panel's assessments as being too light and as too severe in one other case (Cohen's kappa: 0.98). Detailed case description was considered essential to the audit procedure. While aggregated reporting of audit results provides a general understanding of substandard factors in the care provided, feedback of results on an individual practice level led to specific suggestions for improvement (in relation to medical aspects, patient-caregiver relationship and collaboration between caregivers). Lack of anonymity appeared not to be an issue for the caregivers. CONCLUSION: The feedback of perinatal audit results to the caregivers involved as well as discussion of these results led to specific starting points in the areas of collaboration, documentation and policymaking at both individual and institutional level.

Mechanical stimulation of osteopontin mRNA expression and synthesis in bone cell cultures.

We have shown earlier that mechanical stimulation by intermittent hydrostatic compression (IHC) promotes alkaline phosphatase and procollagen type I gene expression in calvarial bone cells. The bone matrix glycoprotein osteopontin (OPN) is considered to be important in bone matrix metabolism and cell-matrix interactions, but its role is unknown. Here we examined the effects of IHC (13 kPa) on OPN mRNA expression and synthesis in primary calvarial cell cultures and the osteoblast-like cell line MC3T3-E1. OPN mRNA expression declined during control culture of primary calvarial cells, but not MC3T3-E1 cells. IHC upregulated OPN mRNA expression in late released osteoblastic cell cultures, but not in early released osteoprogenitor-like cells. Also, in both proliferating and differentiating MC3T3-E1 cells, OPN mRNA expression and synthesis were enhanced by IHC, differentiating cells being more responsive than proliferating cells. These results suggest a role for OPN in the reaction of bone cells to mechanical stimuli. The severe loss of OPN expression in primary bone cells cultured without mechanical stimulation suggests that disuse conditions down-regulate the differentiated osteoblastic phenotype.

Mechanical stimulation by intermittent hydrostatic compression promotes bone-specific gene expression in vitro.

In a previous study of the cellular mechanism underlaying Wolff's law we showed that mechanical stimulation by intermittent hydrostatic compression (IHC) increases bone formation in cultured fetal mouse calvariae compared to non-stimulated cultures. To test whether mechanical stimuli may modulate bone-specific gene expression, we studied the effect of IHC on alkaline phosphatase (AP) expression and enzyme activity as well as collagen and actin mRNA levels in neonatal mouse calvariae and calvarial bone cells. Two cell populations, one resembling osteoprogenitor (OPR) cells and another resembling osteoblasts (OB) were obtained from calvariae by sequential digestion. IHC was applied by intermittently (0.3 Hz) compressing the gas- phase of a closed culture chamber (peak stress 13kPa, peak stress rate 32.5 kPas-1). In control cultures of calvariae as well as OB and OPR cells, AP activity and AP-, collagen-, and actin-mRNA levels all decreased after one or more days, with the exception of OPR cell collagen expression which increased during culture. IHC treatment upregulated AP, collagen and actin expression and AP activity in calvariae and OB cells, but decreased collagen expression in OPR cells. These results suggest that treatment with IHC promotes the osteoblastic phenotype in bone organ cultures and in osteoblasts. Osteoprogenitor cells seem to react somewhat differently to mechanical stress than osteoblasts. The loss of bone-specific gene expression under control culture conditions, in the absence of mechanical stimuli, suggests that the mechanical environment is important in maintaining the differentiated phenotype of bone cells, and that IHC treatment partially restores this environment in bone cell- and organ cultures.

Mechanical loading stimulates the release of transforming growth factor-beta activity by cultured mouse calvariae and periosteal cells.

We have shown earlier that mechanical stimulation by intermittent hydrostatic compression (IHC) inhibits bone resorption and stimulates bone formation in cultured fetal mouse calvariae (Klein-Nulend et al., 1986, Arthritis Rheum., 29: 1002-1009). The production of soluble bone factors by such calvariae is also modified (Klein-Nulend et al., 1993, Cell Tissue Res., 271:513-517). Transforming growth factor-beta (TGF-beta) is an important local regulator of bone metabolism and is produced by osteoblasts. In this study, the release of TGF-beta activity as a result of mechanical stress was examined in organ cultures of neonatal mouse calvariae, in primary cultures of calvariae-derived osteoprogenitor (OPR) cells, and in more differentiated osteoblastic (OB) cells. Whole calvariae and calvariae-derived cells were cultured in the presence or absence of IHC for 1-7 days and medium concentrations of active as well as total TGF-beta were measured using a bioassay. IHC (maximum 13 kPa, maximal pressure rate 32.5 kPa/sec) was generated by intermittently (0.3 Hz) compressing the gas phase above the cultures. We found that mechanical loading by IHC stimulated the release of TGF-beta activity from cultured calvariae by twofold after 1 day. IHC also stimulated the release of TGF-beta activity from calvariae-derived cells after 1 and 3 days. The absolute amounts of TGF-beta activity released were lower in OPR cells than in OB cells, but the stimulatory effect of IHC was greater in OPR cells. Total TGF-beta (active and bound) released into the medium was not affected by IHC. IHC did not change the dry weight of the organ cultures, nor the DNA or protein content of the cell cultures. These data show that mechanical perturbation of bone cells, particularly OPR cells, enhances the activation of released TGF-beta. We conclude that modulation of TGF-beta metabolism may be part of the response of bone tissue to mechanical stress.

Prenatal diagnosis and fetal outcome of cystic adenomatoid malformation of the lung: case report and historical survey.

Diagnostic ultrasound allows prenatal diagnosis of cystic adenomatoid malformation of the lung from the second trimester onwards throughout pregnancy. If the diagnosis is made before the 24th week of pregnancy, counselling often results in termination of pregnancy. Whether or not this attitude is in agreement with good medical practice is discussed on the basis of our own experience and a review of the literature.

Effect of two monophasic oral contraceptives containing gestodene or desogestrel on serum lipoprotein lipid levels.

Forty-nine healthy women aged 20-35 years who had not been pregnant or using an oral contraceptive (OC) for the previous 3 months were randomized into two groups, one group taking an OC containing 75 micrograms gestodene (GTD) and 30 micrograms ethinyl estradiol (EE), and the other group using an OC with 150 micrograms desogestrel (DSG) and 30 micrograms EE. Fasting blood samples were taken before treatment, and after cycles 3 and 6, between the 18th and the 22nd day of the cycle. Blood lipoprotein lipid levels were measured. Serum total cholesterol did not change significantly in both groups. However, in the first three cycles, serum triglyceride increased by 46% and 40% and HDL-cholesterol by 14% and 8% in the GTD and DSG groups, respectively. The serum LDL level decreased by 6.2% and 11.8%, respectively. Between the third and sixth cycle, no further significant changes were observed, nor did these changes differ significantly between the two groups. In conclusion, both OC preparations exerted small and probably favorable effects on serum lipoprotein lipid levels.

Urinary excretion rates of calcium and magnesium in normal and complicated pregnancies.

In this cross-sectional study calcium and magnesium metabolism was investigated in normal pregnancies (n = 34) and pregnancies complicated by either fetal growth retardation of hypertension with or without fetal growth retardation (SGA newborns) (n = 30). Special attention has been given to the renal excretion rates of calcium and magnesium and their relationship to creatinine and sodium clearances. No differences were noted in the third trimester of pregnancy between the normal and complicated pregnancies in calcium or magnesium metabolism except for an increased serum magnesium in the SGA group. Comparing the post-partum period to normal pregnancy the following results were observed: (i) serum ionic calcium levels showed no differences; (ii) urinary calcium excretion was increased as a result of increased calcium clearance. A striking feature was the fact that the fractional calcium clearance was not increased, in contrast to the increase in relative calcium clearance. The observed results can be explained by an increased GFR and a possible dissociation between the sodium and calcium handling in the cortical thick ascending Limb of Henle's Loop.