Bruce A. Buehler

Expression of 72 kDa and 92 kDa type IV collagenases from human giant-cell tumor of bone.

Basement membrane forms widespread barriers to tumor invasion. It has been shown that tumor-secreted, basement membrane-degrading enzymes, namely metalloproteinases (MMPs) play an important role in tumor invasion and metastasis. In this study, we determined the enzymatic activity, content, and mRNA of both the 72 kDa (MMP-2) and 92 kDa (MMP-9) MMPs in primary cultures of human giant-cell tumor of bone (GCT)in vitro and in tissue extracts (in vivo). Gelatin zymography showed the presence of lytic bands at Mr 121000, 92000, and 72000, and these enzymatic activities were inhibited by EDTA, an inhibitor of MMPs. Western blots with antibodies specific for MMP-2 and MMP-9 confirmed the presence of MMP-2 and MMP-9 bothin vitro andin vivo, but GCT cells at late passage showed only MMP-2. Northern blots using labeled cDNA probes specific for these molecules revealed the presence of 3.1 kb transcript for MMP-2 and a 2.9 kb transcript for MMP-9. Using specific antibodies to 72 kDa and 92 kDa type IV collagenases, we studied their cellular distribution by immunohistochemical means. Stronger immunoreactivity was found for 92 kDa type IV collagenase than 72 kDa type IV collagenase in the giant cells. It appears, therefore, that MMP-9 may play an important role in the malignant behavior of GCTs and suggests a potential therapeutic role for protease inhibitors in attempting to minimize the invasive behavior of GCTs.

Vitamin B2: Riboflavin:

Riboflavin (vitamin B 2) is not synthesized by vertebrates and must be supplied by diet. It is found in most foods, with the highest content in dairy products, meat, and dark green vegetables. Deficiency rarely occurs alone where adequate food is available and usually occurs as a multinutrient deficiency. Some conditions and medications can decrease effectiveness or levels of riboflavin. Supplementation is rarely needed to prevent ariboflavinosis when a normal diet is consumed. Supplementation for decreased frequency of migraine headaches and prevention of age related cataracts could be beneficial. There is no known toxicity.

Algorithmic Approach for Methyl-CpG Binding Protein 2 (MECP2) Gene Testing in Patients With Neurodevelopmental Disabilities:

Methyl-CpG binding protein 2 gene (MECP2) testing is indicated for patients with numerous clinical presentations, including Rett syndrome (classic and atypical), unexplained neonatal encephalopathy, Angelman syndrome, nonspecific mental retardation, autism (females), and an X-linked family history of developmental delay. Because of this complexity, a gender-specific approach for comprehensive MECP2 gene testing is described. Briefly, sequencing of exons 1 to 4 of MECP2 is recommended for patients with a Rett syndrome phenotype, unexplained neonatal encephalopathy, an Angelman syndrome phenotype (with negative 15q11-13 analysis), nonspecific mental retardation, or autism (females). Additional testing for large-scale MECP2 deletions is recommended for patients with Rett syndrome or Angelman syndrome phenotypes (with negative 15q11-13 analysis) following negative sequencing. Alternatively, testing for large-scale MECP2 duplications is recommended for males presenting with mental retardation, an X-linked family history of developmental delay, and a significant proportion of previously described clinical features (particularly a history of recurrent respiratory infections).

Complementary and Alternative Medicine (CAM) in Genetics

Complementary and alternative medicine (CAM) is increasingly popular among families. Approximately 50% of children are on herbal preparations, 30% of adolescents take herbal medications, and 70% of adults are involved in some aspect of complementary medicine. The true incidence of CAM use by families who have a family member with special needs or a genetic condition is unknown. Anecdotal evidence suggests that usage may be higher in this group than in the general population. Geneticists and genetic counselors need to be aware of CAM practices with their families and to provide anticipatory guidance.

The Free Radical Theory of Aging and Antioxidant Supplements A Systematic Review

Free radical excess occurs when cells are exposed to reactive oxygen species greater than the amount that can be neutralized by cellular produced antioxidants such as superoxide dismutase. This is termed oxidative stress, which can be caused by excessive energy intake or external pollutants. Excess free radicals are proposed to increase the rate of cell aging, injury, and mutations leading to a shortened cell life span. Vitamins A, C, and E and flavoproteins are supplements that function as free radical scavengers. Antioxidants are present in natural foods but added amounts beyond the diet may detoxify excess free radicals during “oxidative stress.” Antioxidant supplements decrease cellular damage from excess reactive oxygen species but they have not been proven to prolong life span.

Accelerated Linear Growth and Advanced Bone Age in Sotos Syndrome is Not Associated with Abnormalities of Collagen Metabolism

OBJECTIVES To investigate whether the advanced bone age in Sotos syndrome is associated with alterations in type I collagen metabolism in bone. DESIGN AND METHODS The metabolism of collagen was studied by analyzing the production, gene expression and degradation of type I collagen in dermal fibroblast strains from patients with Sotos syndrome and comparing them with fibroblasts from age-matched healthy subjects. Collagen production was determined as collagenase digestible radioactivity and collagen mRNA levels were measured by RT-PCR. Collagen degradation was assessed by specific collagenase assay and gelatin zymography. To determine the structural defects in type I collagen, the newly synthesized proteins were analyzed by SDS-PAGE before and after proteolytic digestion with pepsin. RESULTS In the present study, we have demonstrated that the collagen production, secretion and degradation in Sotos syndrome is comparable to controls. In addition, no qualitative differences in mRNA transcripts for type I collagen were detected between the control and Sotos syndrome fibroblasts. The secretion and intracellular accumulation of procollagen is also comparable to controls. The analysis of both procollagen and collagen on SDS-PAGE did not exhibit any major structural changes as compared with controls. CONCLUSIONS Our results on several aspects of collagen metabolism have demonstrated for the first time that collagen, the most abundant of mammalian proteins and the major constituent of bone, is normal in patients with Sotos syndrome. Therefore, it appears that the advanced bone age and accelerated linear growth seen in patients with Sotos syndrome may not be attributed to inherent abnormalities of collagen metabolism. The etiology and the pathogenesis of Sotos syndrome still remains unclear.