Erythroid potentiating activity of tissue inhibitor of metalloproteinases on the differentiation of erythropoietin-responsive mouse erythroleukemia cell line, ELM-I-1-3, is closely related to its cell growth potentiating activity

The erythroid-potentiating activity (EPA) of the tissue inhibitor of metalloproteinase-1 (TIMP-1) was re-examined using ELM-I-1-3, a mouse erythroleukemia cell line, which responded well to erythropoietin. Depletion of pre-existing TIMP-1 from fetal calf serum in culture medium using monoclonal antibody suppressed erythropoietin-induced differentiation as measured by the induction of hemoglobin, commitment assay and globin mRNAs. The removal of TIMP-1 also suppressed the proliferation of ELM-I-1-3 as measured by cell number and de novo DNA synthesis. These changes were reversed by the addition of purified TIMP-1 to the culture medium. Anti-TIMP-1 antibody also blocked both hexamethylene bisacetamide (HMBA)-induced erythroid differentiation and the proliferation of both ELM-I-1-3 and Friend erythroleukemia cells. Considering previous reports analyzing the chemical induction of Friend mouse erythroleukemia cell differentiation, our results suggest that erythropoietin- or HMBA-induced erythroid differentiation might also be coupled with cell proliferation. Our 3H thymidine-uptake experiment shows that TIMP-1 removal was also effective in the inhibition of cell growth of various other cell lines in addition to erythroleukemia cell lines. These results suggest that EPA action of TIMP-1 on erythroid leukemia cell lines is closely related to its activity to promote the cell growth of various cell lines and cells including erythroleukemia cell lines.     

Mechanisms of expression of pyruvate dehydrogenase deficiency caused by an E1alpha subunit mutation

OBJECTIVE: To characterize the biochemical mechanisms of expression of the pyruvate dehydrogenase (PDH) E1alpha subunit exon 10 R302C missense mutation. BACKGROUND: Mutations in the X-linked E1alpha subunit gene are responsible for most cases of PDH deficiency, an important cause of neurodevelopmental defects and neurodegeneration with primary lactic acidemia. Although the disease shows extreme allelic heterogeneity, the R302C mutation has been defined in several unrelated cases. METHODS: Cell lines expressing selectively either the mutant or wild-type E1alpha alleles against identical genetic backgrounds were generated from the fibroblasts of a female heterozygous for the R302C mutation. Enzyme activity, mRNA, polypeptide expression, and turnover were studied in each. RESULTS: The residual PDH activity was below measurable levels in the cell line (B5) expressing only the mutant allele and normal in the wild-type polypeptide expressing (A10) cell line, confirming that the R302C mutation alone is sufficient to cause a severe PDH deficiency. The mutant polypeptide was less stable than the wild-type polypeptide, but the steady-state level of the mutant E1alpha protein was reduced only two- to threefold. CONCLUSIONS: The primary mechanism of expression of the R302C mutation must be limitation of catalytic efficiency. We speculate that catalysis may be inhibited in the mutant polypeptide because conformational changes are induced near serine 300, a residue that is particularly important as a regulatory phosphorylation site in the wild-type polypeptide.     

Requirement for multiple lymphocyte subsets in protection by a live attenuated vaccine against retroviral infection

STUDY DESIGN: A case report and literature review of thoracic hyperkyphosis deformity secondary to glucocorticoid-induced osteoporosis in Cushing's disease. OBJECTIVES: To identify the pathophysiology of glucocorticoid-induced osteoporosis and to outline the diagnosis and treatment options for a patient with severe spinal deformity secondary to unrecognized excess glucocorticoid activity. SUMMARY OF BACKGROUND DATA: Glucocorticoid-induced osteoporosis is seen in patients exposed to supraphysiologic levels of endogenous or exogenously administered glucocorticoids. In these patients, glucocorticoids act to suppress bone formation and increase bone resorption by indirect and direct effects. These patients have a high prevalence of trabecular bone loss, resulting in much higher rates of vertebral body collapse and pathologic fracture and thus causing an increased propensity toward kyphotic spinal malalignment. METHODS: The literature was reviewed and case reports studied. This case report highlights the pathophysiology of the disease process that caused the spinal deformity and the surgical intervention used to correct the kyphotic deformity after the metabolic problem was resolved. RESULTS: This patient has responded well to treatment and surgical intervention to correct a thoracic hyperkyphotic deformity without complication. CONCLUSIONS: Unrecognized endogenous production of glucocorticoids in Cushing's disease should be considered in young adult patients with progressive osteoporotic spinal deformities.     

The crystal structure of human procathepsin K

Cathepsin K is a cysteine protease present in human osteoclasts that plays an important role in bone resorption. Cathepsin K is synthesized as an inactive proenzyme and activated under conditions of low pH. Autoproteolytic processing of the N-terminal 99 amino acid propeptide produces the active, mature form of cathepsin K. It is presumed that the activation of procathepsin K in vivo occurs in the bone resorption pit, which has a low-pH environment. We have determined the structure of human procathepsin K at 2.8 A resolution. The structure of the mature enzyme domain within procathepsin K is virtually identical to that of mature cathepsin K. The fold of the propeptide of procathepsin K is similar to that observed in procathepsins B and L despite differences in length and sequence. A portion of the propeptide occupies the active site cleft of cathepsin K. Hydrophobic interactions, salt bridges, and hydrogen-bonding interactions are observed in the structure of the propeptide and between the propeptide and the mature enzyme of procathepsin K. These interactions suggest an explanation for the stability of the proenzyme. The structure of procathepsin K contributes to an understanding of the molecular basis of inhibition by the propeptide portion of the molecule and activation of this important member of the cysteine protease family.     

Tinnitus: etiology and management

Tinnitus is the perceived sensation of sound in the absence of acoustic stimulation. Individuals who suffer from it are commonly between the ages of 40 and 80 years. Tinnitus is often classified as objective or subjective, yet the pathophysiologic cause is still unknown. Subjective tinnitus is largely identified with hearing loss. Management of tinnitus is based on an individual approach; there is no single treatment or regimen for it.     

Use of computed radiography in the study of an historic painting

The authors demonstrate the use of radiography in the investigation of an historic painting and describe the potential benefits of computed radiography compared with conventional screen-film radiography. The subject for the comparison was a 16 x 19-foot oil-on-canvas painting, Scipio Africanus Freeing Massiva, by Giovanni Battista Tiepolo. Radiographs of the painting were obtained by using a portable, industrial radiographic unit and both conventional screen-film and photostimulable phosphor plate cassettes. For this investigation, computed radiography had a number of advantages over screen-film radiography, largely due to its wider dynamic range and its capabilities for enhancing the digital images with image processing tools such as magnification, edge enhancement, colorization, and airbrushing. The ability to electronically combine images from the large painting into a single composite image file was extremely valuable, as this technique was much less cumbersome and resulted in much higher quality composite images than could be achieved with conventional radiography. An additional advantage of computed radiography includes the capability to easily archive and transmit these images in a digital format for subsequent review.