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Biomedical subjects

F Shapiro

Publications and source records attributed to F Shapiro.

At least 55 records · Page 3Linked to original sources
PubMed1996

Congenital muscular dystrophy associated with merosin deficiency.

"Classic" congenital muscular dystrophy is a heterogeneous group of disorders, characterized by early-onset muscle weakness and hypotonia, absence of overt cerebral or ocular symptoms, and muscle pathology consistent with a dystrophic process. A subset of patients with congenital muscular dystrophy have recently been found to be deficient in the extracellular matrix protein merosin. Consequently, we reviewed the clinical, pathologic, and immunohistochemical features of 12 patients (six males and six females) with classic congenital muscular dystrophy who have been seen at the Children's Hospital, Boston, over the past 15 years. There was marked clinical heterogeneity within this patient population, with age of independent ambulation ranging from 13 months to 6 years. Immunocytochemical analysis using antibodies to merosin, dystrophin, 43-kDa dystroglycan, adhalin, and laminin was normal in 11 of 12 patients. One patient had markedly abnormal staining for merosin; the majority of fibers were negative, although occasional fibers demonstrated patchy staining. Immunoblot analysis in this patient demonstrated markedly reduced levels of merosin (< 10% compared to controls and other patient), of apparently normal size. Clinically, this patient could be differentiated from the others by a marked elevation of serum creatine kinase (> 1000 U/L) and the presence of early white-matter changes on magnetic resonance imaging. The results of this study support the observation that abnormalities of merosin are present in a subgroup of patients with classic congenital muscular dystrophy. Although marked elevation of serum creatine kinase and white-matter changes on magnetic resonance imaging may serve to distinguish these patients from other patients with congenital muscular dystrophy, there remains a large proportion of patients in whom the underlying pathogenesis remains to be elucidated.

Child↗
PubMed1996

Gadolinium-enhanced MR imaging demonstrates abduction-caused hip ischemia and its reversal in piglets.

PURPOSE: To determine if gadolinium-enhanced MR imaging can detect early reversible ischemia of the capital femoral epiphysis and physis induced by hip hyperabduction in piglets. MATERIALS AND METHODS: Thirteen 1- to 3-week-old piglets were placed in maximal bilateral hip abduction and then studied with dynamic gadolinium-enhanced MR imaging 1-6 hr later to assess ischemia of the 26 femoral heads. The piglets were then allowed to ambulate freely for 1 or 7 days and reimaged in neutral position to assess reperfusion. We evaluated enhancement on MR images and compared them with histologic findings. RESULTS: Decreased or absent enhancement, interpreted as ischemia, developed after maximal hip abduction in all 26 cartilaginous epiphyses and 85% of the 26 physes. The most frequently seen abnormality was a sharply marginated, nonenhancing area in the anterior part of the femoral head. A smaller area of decreased enhancement developed in the posterior part of the femoral head adjacent to the acetabular rim. The secondary center of ossification was ischemic in 10 (56%) of the 18 hips after 1 hr of abduction and in all 8 hips after 4 or 6 hr (p = .02). The overall severity of ischemia was greater with longer abduction times (p < .001) and greater degrees of abduction (p < .01). Reperfusion was complete in two (17%) of the 12 hips after 1 day of ambulation and in all 10 (100%) after 1 week of ambulation. CONCLUSION: Enhanced MR imaging detects early ischemia of the epiphyseal and physeal cartilage and the epiphyseal marrow. In piglets, ischemia due to maximal abduction is reversible if corrected within 6 hr.

Age Factors↗
PubMed1995

Human bone marrow microvascular endothelial cells support long-term proliferation and differentiation of myeloid and megakaryocytic progenitors.

Endothelial cells are a major component of the bone marrow (BM) microenvironment that regulate the trafficking and homing of hematopoietic progenitor and stem cells. In this paper, we provide evidence that BM endothelial cells (BMECs) also support multilineage hematopoiesis by elaboration of soluble cytokines. Hematopoietic progenitor cells incubated in direct contact with BMEC monolayers, or physically separated by microporous membrane, expanded five-fold to sevenfold at 7 days, in the absence of exogenous cytokines. Flow cytometric analysis of proliferating progenitor cells grown in the presence of BMEC monolayers showed that by day 14 of coculture, 70% to 80% of hematopoietic cells were myeloid, expressing CD15 or CD14, and 14% to 19% were megakaryocytic, expressing GPIIb/IIIa or GPIb. CD34+ cells derived from umbilical cord blood, cultured in the upper chamber of transwell culture plates, as well as the cells grown in direct contact with BMEC monolayers, generated progenitors for up to 70 days. Unstimulated BMEC monolayers constitutively produce interleukin-6, Kit-ligand, granulocyte colony-stimulating factor, and granulocyte macrophage colony-stimulating factor. These data suggest that BMEC regulate proliferation of hematopoietic progenitor cells and long-term culture initiating cells by elaboration of lineage-specific cytokines.

Antigens, CD↗
PubMed1995

Transmission electron microscopic demonstration of vimentin in rat osteoblast and osteocyte cell bodies and processes using the immunogold technique.

BACKGROUND: The immunogold labeling technique and transmission electron microscopy were used to demonstrate the expression and position of the intermediate filament vimentin in rat osteoblast and osteocyte cell bodies and cell processes. Conventional light and transmission electron microscopic studies of bone cells demonstrated adjacent cell linkage to be mediated by osteoblast and osteocyte processes present within the canalicular system traversing the bone matrix. The cell processes were filled with densely packed filaments, many of which have been shown previously to be actin microfilaments. The appearance, however, of 10 nm diameter filaments in some cell processes and the fact the intermediate filament vimentin has been defined in many cells of mesenchymal origin raised the possibility that some of these filaments might be vimentin. The ultrastructural colloidal gold immunochemical technique allowed for demonstration in situ of the expression of vimentin filaments plus accurate definition of their position. METHODS: The studies were performed in newborn rat femoral and tibial diaphyseal cortical bone and in 1-week-old repair bone from 2.4 mm diameter defects made through the lateral cortex in 6-week-old rat femurs and tibias. The bone tissues for the immunochemical study were fixed in 1% glutaraldehyde, 4% paraformaldehyde, and 0.1 M phosphate buffer (pH 7.4) for 2 days. Decalcification was performed in 6% EDTA for 2-3 days. Infiltration involved use of Lowicryl resin K4M, and the embedding and curing processes were performed in a cryostat with temperatures -30 degrees C. An antivimentin monoclonal antibody was used for labeling using the postembedding technique. Effective antibody dilutions ranged from 1:10 to 1:200, with the dilutions of 1:25 and 1:100 showing the best combination of filament labeling with the least matrix background. The grids were exposed to 10 nanometer gold colloid conjugated goat anti-mouse IgM for demonstration of binding. RESULTS: Vimentin immunolabeling was defined clearly in relation to filaments within the osteoblast and osteocyte cell body cytoplasm, throughout the entire length of the osteoblast and osteocyte cell processes, and in close relationship to the intracellular gap junctions which were present within the cell processes both close to the cell bodies and within the canaliculi well away from them. CONCLUSIONS: Immunogold labeling demonstrates the presence of the intermediate filament vimentin in osteoblast and osteocyte cell bodies and processes of rat bone. Vimentin distribution is not concentrated to specific areas, is present throughout the extent of the bodies and processes, and is seen immediately adjacent to gap junctions.

Age Factors↗
PubMed1995

Gadolinium-enhanced MR imaging demonstrates abduction-caused hip ischemia and its reversal in piglets.

Purpose. To determine whether gadolinium-enhanced MR imaging can detect early reversible ischemia of the capital femoral epiphysis and physis induced by hip hyperabduction in piglets. Materials and methods. Thirteen 1- to 3-week-old piglets were placed in maximal abduction of both hips and studied with dynamic gadolinium-enhanced MR imaging 1-6 h later to assess ischemia of the 26 femoral heads. They were then allowed to ambulate freely for 1 or 7 days, and reimaged in neutral position to assess reperfusion. Enhancement was evaluated on MR images and compared with histologic findings.Results. Ischemia after hyperabduction developed in all 26 cartilaginous epiphyses and in 85 % of the physes. The most frequent abnormality was a sharply marginated nonenhancing area in the anterior part of the femoral head. A smaller area of ischemia developed in the posterior part of the femoral head, adjacent to the acetabular rim. The secondary center of ossification was ischemic in 56 % of the hips after 1 h of abduction and in all hips after 4 or 6 h (p = 0.02). The overall severity of ischemia was greater with increasing abduction time (p < 0.001) and increasing degree of abduction (p < 0.01). There was partial reperfusion in 83 % of the hips after 1 day of ambulation and complete reperfusion in all 26 hips (100 %) after 1 week.Conclusion. Enhanced MRI detects early ischemia of the epiphyseal and physeal cartilage and the epiphyseal marrow. In piglets, ischemia due to hyperabduction is reversible if corrected within 6 h.

Animals↗
PubMed1995

Characterization of hematopoietic cells arising on the textured surface of left ventricular assist devices.

BACKGROUND: Textured biomaterial surfaces in implantable left ventricular assist devices induce development of a nonthrombotic neointimal surface and allow elimination of anticoagulation therapy in device recipients. Characterization of the hematopoietic cells formed within the neointimal surfaces of these devices will contribute to our understanding of this unique neointima. METHODS: The blood-contacting surface of seven ThermoCardiosystems left ventricular assist devices was removed, washed with phosphate-buffered saline solution, and digested with 0.1% collagenase for 15 to 20 minutes. The hematopoietic cells released from the explants were isolated and analyzed by flow cytometry and immuno-histochemical staining. RESULTS: More than 80% +/- 6% of hematopoietic cells isolated in this fashion are of myelomonocytic origin and express CD14, CD15, and CD33 surface molecules. Four percent of cells express the CD34 surface marker, which suggests that the neointima is colonized by pluripotent hematopoietic stem cells. Continuous culture of these hematopoietic cells in the presence of the cytokines interleukin-3, c-kit ligand, granulocyte colony-stimulating factor resulted in tenfold expansion by day 7 and 25-fold expansion by day 14. CONCLUSIONS: Pluripotent hematopoietic cells with a high proliferative capacity colonize textured surfaces of left ventricular assist devices and may contribute to the development of a biologically nonthrombogenic neointima.

Antigens, CD↗
PubMed1995

Stunting syndrome in broilers: physical, physiological, and behavioral aspects.

The effect of stunting syndrome (SS) on young broilers induced by orally administering an inoculum prepared from intestines of SS-affected broiler chicks was studied in two experiments. Depression of growth, feed intake, and feed utilization, respectively, was negatively related to the age of inoculation, i.e., highest at posthatch inoculation (63, 57, and 61%), and intermediate at the ages of 3 (42, 45, and 50%) and 7 d (29, 34, and 53%), whereas at the age of 14 d inoculation was ineffectual and the inoculated chicks performed similarly to the naive controls. Eating behavior was determined by periodically recording the number of chicks in each treatment group exhibiting this behavior, i.e., pecking mash in the feeder or pecking the litter. Eating activity was much higher in inoculated chicks (about 20 to 24%) than in the naive controls (6 to 12%) and as with performance negatively related to the age of inoculation. In chicks inoculated at the age of 14 d, eating activity was quite similar to that of the naive control chicks. Noninoculated chickens raised in the same room as their inoculated counterparts were infected by the disease agents. In some respects the consequences of horizontal infection were similar to those observed in inoculated chicks, i.e., depressed feed intake, growth, and feed utilization and a heavier heart, crop, proventriculus, gizzard, intestine, and gastrointestinal contents. In contrast, the activities of the pancreatic digestive enzymes were more similar to those of the naive controls than to those of the inoculated groups. At the age of 14 d, activities of amylase, trypsin, chymotrypsin, and lipase in the pancreas were lower in the inoculated than in the naive control birds. At the age of 21 d, the results were reversed and activity in the inoculated birds was higher than in the naive control birds. During both periods, the activity of pancreatic lipase was higher in the naive controls than in the inoculated birds. The hyperactivity estimated via a determination of eating activity suggests that SS is an affliction not only of digestion but also of metabolism.

Aging↗
PubMed1995

Stunting syndrome in broilers: effect of age and exogenous amylase and protease on performance, development of the digestive tract, digestive enzyme activity, and apparent digestibility.

Day-old male, meat-type chicks raised in brooder batteries were infected by orally administering an inoculum prepared from intestines of broiler chicks infected with stunting syndrome (SS). Naive controls were kept in a parallel room. The chicks were fed a commercial starter diet supplemented with two levels of enzyme preparations to 14 d of age. The experiment was continued to the age of 6 wk in order to estimate compensatory feed intake and growth. In a parallel study, digestibility of the feed was determined from 1 to 3 wk of age with control or inoculated chicks. The enzymes amylase and proteases were produced by Bacillus subtilis and Penicillium emersonii. Enzyme supplementation had no effect on feed intake, growth, or feed utilization, or on digestibility of fat, starch, protein, or energy. Because enzyme supplementation did not consistently affect performance of chicks and no interactions were observed between enzyme supplementation and infection status, data are presented for effects of infection only. Inoculation of SS-infective material reduced performance to 4 wk. Compensatory growth and feed intake were observed from the age of 4 wk onward. At the age of 6 wk the slight retardation of the inoculated chicks was not significant. On Week 1, retention of fat, starch, protein, and energy was significantly depressed in the inoculated chicks. At the age of 2 wk, retention of starch was not depressed, and at the age of 3 wk, the only consistent depression was that observed for fat. The proventriculus weight and content were consistently higher in inoculated chicks, as were the small intestine and intestinal content. The pH of the gizzard content was higher, and that of the small intestine content was lower, in the inoculated birds than in their control counterparts. Stunting syndrome infection was accompanied by a significant depression of trypsin activity in the pancreas at the age of 1 and 2 wk. At these periods, amylase and chymotrypsin were not affected. At 6 wk of age, the activities of amylase, trypsin, and chymotrypsin in the pancreas were higher in the inoculated than in the control birds. In the intestinal chime, amylase, trypsin, and chymotrypsin activities were lower in the inoculated birds on Week 1 and 2 (NS for amylase on Week 1). On Week 6, the activity of all enzymes assayed was higher in the inoculated birds (NS for amylase). It is suggested that the main factors depressing feed intake and growth in SS-infected birds are most probably beyond those of digestion.

Aging↗
PubMed1994

Optimization of conditions for ex vivo expansion of CD34+ cells from patients with stage IV breast cancer.

Multiple cycles of high-dose chemotherapy can be hematologically supported by repeated administration of peripheral blood progenitors obtained after mobilization using cytokine alone or in combination with chemotherapy. We have explored the quality of such cells and their potential to undergo ex vivo expansion. Twenty-five leukapheresis samples from 19 patients who had received extensive prior chemotherapy for stage IV breast cancer were subjected to CD34+ cell selection using immunoaffinity columns of immunomagnetic bead separation. Cells were cultured in suspension in the presence of c-kit ligand, interleukin-3, interleukin-6, erythropoietin, and granulocyte colony-stimulating factor. Ten experiments were performed using weekly exchange of media and cytokines (Delta assay). Median myeloid and erythroid progenitors expanded 15-fold at 7 days (range, 7 to 43), 40-fold at 14 days (range, 18 to 470), 46-fold at 21 days (range, 0 to 118), and 21-fold at 28 days (range, 0 to 61). In a system using gas-permeable bags without exchange of media or cytokine, median progenitors expanded 13-fold at 7 days (range, 7 to 36), 14-fold at 10 days (range, 4 to 61), 14-fold at 12 days (range, 3 to 46), and 10-fold at 14 days (range, 1 to 35). Progenitor expansion less than 10-fold occurred in 8% of experiments at day 7, in 17% at day 10, in 43% at day 12, and in 50% at day 14. When autologous plasma, autologous plasma processed (removal of cryoprecipitate, centrifugation, then filtration), or human serum were substituted for 20% fetal calf serum, the ratio of progenitor expansion at 7 days relative to 20% fetal calf serum for 10% human serum, 20% human serum, and 1% autologous plasma processed was 1.01 (range, 0.62 to 1.33), 0.88 (range, 0.61 to 1.20), and 0.96 (range, 0.55 to 1.64), respectively. These findings support the feasibility of ex vivo expansion in a system free of nonhuman proteins of CD34(+)-derived progenitors obtained from the peripheral blood of patients who have received prior chemotherapy.

Animals↗
PubMed1994

Isolation and characterization of human bone marrow microvascular endothelial cells: hematopoietic progenitor cell adhesion.

To examine potential mechanisms by which hematopoiesis may be regulated by endothelial cells within the bone marrow (BM) microenvironment, we have devised a technique for the in vitro study of the interaction of human BM microvascular endothelial cells (BMEC) with hematopoietic cells. Microvessels isolated by collagenase digestion of spicules obtained from filtered BM aspirate were plated on gelatin-coated plastic dishes, and colonies of endothelial cells grown from microvessel explants were further purified by Ulex europaeus lectin affinity separation. BMEC monolayers isolated by this technique grew in typical cobblestone fashion, stained positively with antibody to factor VIII/von Willebrand factor, and incorporated acetylated LDL. Immunohistochemical studies showed that BM microvessels and BMEC monolayers express CD34, PECAM, and thrombospondin. Incubation of resting BMEC with BM mononuclear hematopoietic cells resulted in the selective adhesion of relatively large numbers of CD34+ progenitor cells and megakaryocytes. The binding of purified BM-derived CD34+ progenitor cells to BMEC was dependent on divalent cations and was partially blocked by antibodies to CD34. IL-1 beta treatment of BMEC monolayers resulted in an increase of CD34+ progenitor cell adhesion by mechanisms independent of CD34 or divalent cations. BMEC exhibit specific affinity for CD34+ progenitor cells and megakaryocytes, suggesting that the BM microvasculature may play a role in regulating the trafficking, proliferation, and differentiation of lineage specific hematopoietic elements, and possibly of pluripotent stem cells within the CD34+ population.

Antigens, CD↗
PubMed1994

Abnormality of type IX collagen in a patient with diastrophic dysplasia.

There is growing evidence that a spectrum of chondrodysplasias are caused by mutations in the gene coding for type II collagen. The basic molecular defect in diastrophic dysplasia has not been defined, but it appears not to be in collagen type II. Cartilage contains other tissue-specific collagens, types IX, X, and XI, but no mutations have yet been found in their genes in clinical disease. Type IX collagen is hypothesized to play a role in the regulation of type II collagen fibril organization and structure in cartilage extracellular matrix. In this study, we have examined iliac crest growth cartilage from a patient with diastrophic dysplasia. Although collagen fibrils were markedly increased in diameter on transmission electron microscopy, type II collagen appeared to be normal biochemically. Type XI collagen was also normal. However, type IX collagen appeared abnormal on sodium dodecyl sulfate polyacrylamide gel electrophoresis with a pronounced excess of the COL1 domain of the molecule in pepsin extracts. The findings point to an abnormality in structure or metabolism of type IX collagen in diastrophic dysplasia.

Amino Acid Sequence↗
PubMed1994

Regulation and function of hematopoietic stem cells.

Hematopoietic stem cells, defined as cells with extensive self-renewal and pluripotent differentiation potential, represent a minor population of adult bone marrow (< 1 in 10(4) to 10(5) nucleated marrow cells). Recent advances in cell surface phenotype analysis and separation technology have permitted enrichment of hematopoietic stem cells. Numerous cytokines have been identified that interact additively or synergistically with hematopoietic stem cells to regulate their self-renewal and differentiation. Qualitative and quantitative analysis of hematopoietic stem cells has increasing clinical importance in areas of stem cell-toxic therapy, allogeneic or autologous hematopoietic transplantation, and stem cell gene therapy for genetic or acquired diseases.

Animals↗
PubMed1993

In situ hybridization studies suggest a role for the basic region-leucine zipper protein hXBP-1 in exocrine gland and skeletal development during mouse embryogenesis.

The spatial and temporal distribution of transcripts for the TRE/CRE-binding basic region-leucine zipper protein hXBP-1 was determined by in situ hybridization. Analysis of embryos from day 10.5 to 18.5 pc revealed high level expression of hXBP-1 RNA in two developing organ systems: 1) in bone and cartilage cells of the developing skeleton and toothbuds, and 2) in exocrine glands including the pancreas and the submandibular and salivary glands. High level expression was also found in whisker follicles and in selected cells in brown adipose tissue. In the developing skeleton, hXBP-1 RNA was expressed starting on day 11.5 pc in osteoblasts of newly formed intramembranous bone. Thereafter, hXBP-1 was expressed in both osteoblasts and preosteoblasts in bone formed directly by intramembranous formation as well as in bone formed during endochondral ossification. The most intense signal was observed in preosteoblasts and osteoblasts of newly forming bone. At day 11.5 pc low level hXBP-1 expression was also observed in matrix secreting chondroblasts of bones which are formed initially of cartilage, at the stage where they consist entirely of cartilage. Signal was also present in matrix producing chondroblasts of the mature zone of the growth region during endochondral ossification although at significantly lower level than in osteoblasts. hXBP-1 is thus the first transcription factor described, to our knowledge, whose level of expression is modulated during the osteoblast developmental sequence in vivo. The pattern of expression of hXBP-1 in the developing skeleton was found to be very similar to that of the genes encoding the tissue inhibitor of metalloproteinase and alkaline phosphatase throughout development. These observations suggest that hXBP-1 may play a role in regulating the expression of tissue specific genes (TIMP, osteonectin, osteopontin, osteocalcin) expressed in osteoblasts. It is intriguing that the promoter regions of several such genes contain potential hXBP-1 binding sites.

Adipose Tissue↗
PubMed1993

Cell origin and differentiation in the repair of full-thickness defects of articular cartilage.

The origin and differentiation of cells in the repair of three-millimeter-diameter, cylindrical, full-thickness drilled defects of articular cartilage were studied histologically in New Zealand White rabbits. The animals were allowed to move freely after the operation. Three hundred and sixty-four individual defects from 122 animals were examined as long as forty-eight weeks postoperatively. In the first few days, fibrinous arcades were established across the defect, from surface edge to surface edge, and this served to orient mesenchymal cell ingrowth along the long axes. The first evidence of synthesis of a cartilage extracellular matrix, as defined by safranin-O staining, appeared at ten days. At two weeks, cartilage was present immediately beneath the surface of collagenous tissue that was rich in flattened fibrocartilaginous cells in virtually all specimens. At three weeks, the sites of almost all of the defects had a well demarcated layer of cartilage containing chondrocytes. An essentially complete repopulation of the defects occurred at six, eight, ten, and twelve weeks, with progressive differentiation of cells to chondroblasts, chondrocytes, and osteoblasts and synthesis of cartilage and bone matrices in their appropriate locations. At twenty-four weeks, both the tidemark and the compact lamellar subchondral bone plate had been re-established. The cancellous woven bone that had formed initially in the depths of the defect was replaced by lamellar, coarse cancellous bone. Autoradiography after labeling with 3H-thymidine and 3H-cytidine demonstrated that chondrocytes from the residual adjacent articular cartilage did not participate in the repopulation of the defect. The repair was mediated wholly by the proliferation and differentiation of mesenchymal cells of the marrow. Intra-articular injections of 3H-thymidine seven days after the operation clearly labeled this mesenchymal cell pool. The label, initially taken up by undifferentiated mesenchymal cells, progressively appeared in fibroblasts, osteoblasts, articular chondroblasts, and chondrocytes, indicating their origin from the primitive mesenchymal cells of the marrow. Early traces of degeneration of the cartilage matrix were seen in many defects at twelve to twenty weeks, with the prevalence and intensity of the degeneration increasing at twenty-four, thirty-six, and forty-eight weeks. Polarized light microscopy demonstrated failure of the newly synthesized repair matrix to become adherent to, and integrated with, the cartilage immediately adjacent to the drill-hole, even when light microscopy had shown apparent continuity of the tissue. In many instances, a clear gap was seen between repair and residual cartilage.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗