Biomedical subjects
Publications and source records attributed to R Lindeman.
Inhibitors are an uncommon complication of mild haemophilia A but represent a severe disease, typically with high titre inhibitors and an associated high rate of bleeding. We present data from three patients with MHAI who were successfully treated with Rituximab alone and unequivocally prove that such inhibitors respond to this agent. A treatment protocol is suggested.
Gene therapy has been investigated in many aspects of plastic and reconstructive surgery. These areas ultimately involve various forms of tissue healing and the manipulation of bony and soft tissues to reconstruct defects secondary to neoplastic and congenital disorders and trauma. Most research has been limited to animal studies with the exception of clinical trials on the use of gene therapy in lower leg ulcer healing and as an adjunct to advanced recurrent squamous cell carcinoma of the head and neck. Overall, these preliminary studies have produced optimistic results. With the development of more efficient and safer delivery systems, the application of gene therapy in plastic surgery could become more widespread, especially in combination with tissue engineering technology.
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BACKGROUND: Pancreatic insulin-producing g-cells are permanently destroyed in Type I diabetic patients, leading to hypoglycemica. Various somatic cells have been studied for their ability to deliver insulin as an alternative source of pancreatic g-cells. We investigated the potential of human BM progenitor cells for this purpose. METHODS: Two BM-derived hematopoietic cell lines, Tf-1 (CD34+) and K562 (CD34m) cell and primary human BM stromal cells were transduced with the human preproinsulin cDNA, and the ability of these cells to synthesize, store and release insulin was analyzed. RESULTS: All cells produce and released (pro)insulin at 116-295 wU/10(6) cells/day respectively. No storage of insulin was detected in either cell line or in stromal cells. DISCUSSION: We conclude that human BM-derived progenitor cells can be induced to produce and release basal levels of (pro)insulin.
Gene therapy with the complementary DNA (cDNA) of the angiogenic cytokine vascular endothelial growth factor (VEGF) has emerged as a promising strategy in the treatment of myocardial and lower-limb ischaemia. The objective of this study was to determine whether these principles could be applied to a recognised model of skin-flap ischaemia. Plasmid vectors including the cDNA of green fluorescent protein (GFP) and one of three VEGF isoforms (A165, B167 or B186) were constructed, and their base sequences confirmed. GFP expression was used as a marker of successful in vitro transfection of human endothelial cells with each plasmid. The plasmids were then administered subcutaneously to rat abdominal skin flaps surgically rendered ischaemic, and the percentage of viable tissue was assessed at 1 week. Angiograms of the flaps and histological preparations of flap tissue were assessed for evidence of angiogenesis. The survival of flaps treated with VEGF A165 or B167 cDNA was significantly greater than that of controls (P < 0.05). The survival of flaps treated with VEGF B186 cDNA was greater than that of controls, but statistical significance was not reached. Angiograms and microvessel density counts failed to produce evidence of angiogenesis. With improved delivery strategies, VEGF may have a role in the management of surgical ischaemia.
Acute myeloid leukaemia (AML) blasts rarely express the B7 family of co-stimulatory molecules and do not elicit a clinically significant autologous T-lymphocyte anti-tumour response. The aim of this study was the in vitro modification of AML blasts to an antigen-presenting cell phenotype characterised by upregulated expression of the co-stimulatory molecule CD80 (B7-1). Circulating AML cells were induced to undergo partial differentiation in culture with the cytokines IL-3, IL-6 and GM-CSF; they exhibited variable upregulation of CD80 and continued to express MHC class I and II. These cells remained viable to day 20, in contrast with normal peripheral blood mononuclear cells (PBMNC), which did not survive under the culture conditions. In contrast to unmanipulated blasts, cultured leukaemic cells expressed B7-1. Where initial cytogenetic abnormalities were present, they were also seen in flow-sorted CD80-expressing cells after culture in cytokines, indicating their malignant origin. The immunogenic potential of these cultured cells was highlighted by allogeneic and autologous mixed lymphocyte reactions, in which both differentiated, but not unmanipulated, blasts produced expansion of T-lymphocyte numbers. Autologous cytotoxic T-lymphocyte (CTL) assays indicated specific killing of B7-1+ leukaemic cells, which was greatly enhanced after priming of the T-lymphocytes by B7-1+ blasts prior to the CTL assay, then enabling the CTL to lyse both unmanipulated and differentiated leukaemic cells.
The use of exogenous agents to stimulate the growth of new blood vessels into ischaemic tissue is a potentially revolutionary therapy in a wide variety of clinical specialties. Therapeutic angiogenesis research has been mostly confined to ischaemia of the heart and the lower limb. There has been relatively little research into the potential applications of the technique to plastic, reconstructive and burns surgery. In this paper, relevant published work is reviewed and potential applications of therapeutic angiogenesis to our specialty are considered.
Human long-term bone marrow stromal cultures (LTBMC) have been transduced to express the immunostimulatory cytokine interleukin-2 (IL-2). LTBMC were established from normal bone marrow and characterized by gross morphology, immunohistochemistry, immunophenotyping, and immunofluorescent staining. A stromal marrow cell (SMC) population expressing the markers CD68, CD29, CD13, and CD54 was identified. These cells were successfully transduced with an adenoviral vector containing the human IL-2 gene. Production of IL-2 was demonstrated for at least 13 days post-transduction and was maximal on day 10 at 19,350 pg (or 38 IU) of IL-2/10(6) cells/24 h. IL-2 expressed by the transduced SMC was demonstrated to be biologically active, resulting in the proliferation of autologous T lymphocytes in a mixed lymphocyte reaction. Autologous human LTBMC can be obtained and manipulated with relative ease and may be a useful vehicle for sustained local cytokine delivery.
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Severe combined immunodeficiency is a heterogenous syndrome of varied genetic origins of which the X-linked type is the commonest (XSCID). The most sensitive method for diagnosis of XSCID in the absence of X-linked inheritance pattern is by mutation analysis. In this report we have performed mutation analysis in 13 unrelated boys transplanted (BMT) for SCID without a known cause to determine the frequency of XSCID. Five boys had an affected male relative. We also assessed the utility of hair roots for children without pre-transplant blood stored for mutation analysis since donor genotype was expressed in peripheral blood post BMT. Screening was performed by analysis of single-strand conformational polymorphism (SSCP) followed by sequencing of candidate exons. Mutations were found in 11 cases, of which six were sporadic, and maternal mosaicism was found in one family. Three mothers of the six sporadic cases were identified as carriers. The majority (6/8) of boys with SCID had gammac deficiency despite the absence of X-linked inheritance pattern. The significant frequency of de novo mutations and the occurrence of maternal germline mosaicism highlights the importance of mutation analysis. The strategy of using DNA from hair roots was particularly valuable where no pre-transplant blood was stored. Characterization of the mutations will also enable research into the correction of these genetic defects.
In the context of medicine cutting is one of the most important operations. Many surgical tasks start with an incision, allowing the surgeon to access the region of interest, using either conventional or minimal invasive surgery techniques. The work presented herein should be seen in the context of surgical training and preoperative planning using computer assisted medical simulation systems. The main result of this work is the proposed cutting algorithm based on predefined templates. To integrate this approach in the existing surgical training simulator and to handle additional tasks like soft tissue deformation, needed by the medical environment, a generic simulation framework was developed. This framework is able to utilize given multiprocessor- and network-environments to achieve the desired realtime interactivity.
We performed gamma c gene analysis of a boy with severe combined immunodeficiency whose brother died of the same condition. A base pair deletion in exon 6 was found, which was absent in his mother, indicating maternal mosaicism with important implications for genetic counseling and demonstrating the importance of mutation analysis in boys with severe combined immunodeficiency.
OBJECTIVE: Carrier identification in X-linked immunodeficiency disorders can be based on the demonstration of non-random X inactivation (NRXI) in affected blood cell lineages when growth is impaired in cells expressing the abnormal gene. We examined the utility of seeking evidence of NRXI to test the carrier status of women in families affected by X-linked severe combined immunodeficiency (XSCID) and X-linked hypogammaglobulinaemia (XLH), to identify as carriers the mothers of boys with SCID or hypogammaglobulinaemia whose phenotype suggested X-linkage and to infer X-linkage in boys with SCID or hypogammaglobulinaemia whose disease was not clearly X-linked on the basis either of family history or clinical and immunological characteristics. METHODOLOGY: A polymerase chain reaction-based method was used to amplify a polymorphic CAG repeat in the first exon of the androgen receptor gene after selective digestion of the active X chromosome with a methylation-sensitive enzyme, HpaII to distinguish between the paternal and maternal alleles and to identify their methylation status. RESULTS: Heterozygosity was found in 24 of 31 female subjects (77%). As anticipated, NRXI could be demonstrated in all lymphoid cells studied from obligate carriers of XSCID and an obligate carrier of XLH but not on a carrier of X-linked immunodeficiency with hyper-IgM. The finding of NRXI in the mother of a boy with a SCID variant showed her to be a carrier of XSCID and establishes that her son has XSCID, not otherwise evident from available data. CONCLUSIONS: This PCR assay provides a rapid method for carrier detection of X-linked immunodeficiencies, and has allowed us to expand the phenotype of XSCID
This paper describes a Virtual Environment system designed to aid in training interventional radiologists in inferior vena cava filter placement. It is being developed as part of a VE simulator for a number of surgical and interventional radiology procedures at the Laboratory for Advanced Computer Applications in Medicine at the George Washington University. In this procedure a filter is placed in the inferior vena cava to prevent blood clots from the lower portion of the body from reaching the lungs and causing a pulmonary embolus. The simulation is designed to provide both tutorial and testing modes for the filter placement procedure.
Heparin-induced thrombocytopenia (HIT) is mediated by a heparin-dependent antibody/platelet factor 4/heparin complex binding to platelets via the Fc gamma receptor (type IIA). A single base polymorphism at position 131 of Fc gamma RIIA changes the native arginine to histidine. In the presence of murine monoclonal IgG1 the former phenotype (Fc gamma RIIAArg131) is functionally characterized by strong platelet aggregation (high responder) and the latter (Fc gamma RIIAHis131) by poor aggregation (low responder). In the presence of human IgG2 the opposite response is observed. It has recently been shown that the heparin-dependent antibody is predominantly of this subclass. We hypothesize that a relationship exists between Fc gamma RIIAHis131 and the development of HIT. We studied 24 normal individuals and 20 HIT patients using VM58, a murine monoclonal IgG1, to characterize the phenotype by platelet aggregrometry, and PCR products, amplified with primers bordering the Fc gamma RIIA polymorphism and hybridized with oligonucleotide probes specific for the single base mutation, to determine the genotype. The distribution of phenotypes and genotypes in the two populations differed, with a greater prevalence of the Fc gamma RIIAHis131 allele in the HIT patient population. Homozygous Fc gamma RIIAArg131 individuals were absent from this group. We conclude that the presence of the Fc gamma RIIAHis131 allele is associated with a predisposition to HIT.
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An extensive set of chromosome 15 DNA polymorphisms and densitometric analysis with four markers mapping to the Prader-Willi chromosome region (PWCR) of chromosome 15 have been used to characterize a cohort of 30 subjects with classical Prader-Willi syndrome (PWS). Molecular analysis enabled the classification of the PWS subjects into four groups: (A) 18 subjects (60%) had deletions of paternal 15q11-13 involving a common set of DNA markers. Two subjects had differently sized deletions, one larger and one smaller than the other cases. (B) Eight (27%) had maternal uniparental disomy for chromosome 15. (C) One (3%) had a marker chromosome carrying an extra copy of the PWCR. The marker chromosome was demonstrated to be of paternal origin and the two intact chromosomes were maternally derived. This case represents an apparent exception to the generally held view that PWS is associated with an absence of paternally inherited gene(s) located in the PWCR. (D) The remaining three cases (10%) had none of the above abnormalities. This last subgroup of patients has not previously been well characterized but could represent limited deletions not detectable with the markers used or abnormalities in the imprinting process. These cases represent potentially valuable resources to elucidate more precisely the fundamental disorders responsible for PWS.
The Angelman syndrome (AS) and Prader-Willi syndrome (PWS) loci have been mapped to chromosome 15q11-q13. Chromosomal deletions of differing parental origin in the two syndromes have been interpreted as being due to genetic imprinting. Molecular analysis of patients with varying deletions has localized the AS locus to the interval between D15S113 and GABRB3 and the PWS locus between D15S13 and D15S113. In the present study, DNA cloning and physical mapping techniques have been used to characterize the AS/PWS chromosome region in the vicinity of D15S10, a locus that is telomeric to D15S113 and centromeric to GABRB3. A CpG island near TD3-21 at D15S10 has been cloned, allowing the identification of a widely expressed 4.5-kb transcript and providing a novel DNA marker, OP3, at this locus. OP3 and TD3-21 have been used to construct a long-range physical map extending over approximately 2800 kb. Clusters of rare-cutting restriction sites on this map locate four other CpG islands. Since these CpG islands lie within the minimum deletion intervals for AS and PWS, they mark the possible locations of candidate genes for the two syndromes.