Biomedical subjects

N S Gibran

Publications and source records attributed to N S Gibran.

At least 19 recordsLinked to original sources

PubMed2001

Color subtractive-computer-assisted image analysis for quantification of cutaneous nerves in a diabetic mouse model.

Immunohistochemistry (IHC) is a valuable tool for labeling structures in tissue samples. Quantification of immunolabeled structures using traditional approaches has proved to be difficult. Manual counts of IHC-stained structures are inherently biased, require multiple observers, and generate qualitative data. Stereological methods provide accurate quantification but are complex and labor-intensive when staining must be compared among large numbers of samples. In an effort to quickly, objectively, and reproducibly quantify cutaneous innervation in a large number of counterstained tissue sections, we developed a color subtractive-computer-assisted image analysis (CS-CAIA) system. To develop and test the CS-CAIA method, tissue sections of diabetic (db/db) mouse skin and their wild-type (db/-) littermates were stained by IHC for the neural marker PGP 9.5. The brown-red PGP 9.5 peroxidase stain was colorimetrically isolated through a scripted process of color background removal. The remaining stain was thresholded and binarized for computer determination of nerve profile counts (number of stained regions), area fraction (total area of nerve profiles per unit area of tissue), and area density (total number of nerve profiles per unit area of tissue). Using CS-CAIA, epidermal nerve profile counts, area fraction, and area density were significantly lower in db/db compared to db/- mice.

Animals↗

PubMed2000

Nerve distribution in hemangiomas depends on the proliferative state of the microvasculature.

Hemangiomas appear at birth and undergo gradual regression within several years. Recent published studies have documented increased nerve numbers in port-wine stains and intramuscular vascular tumors. The aim of this study was to establish a relationship between angiogenesis and nerve growth in lesions that undergo neovascular proliferation followed by vessel involution. Twenty-two hemangiomas and arteriovenous malformations were studied using indirect immunocytochemistry with antibodies against the nerve markers protein gene product 9.5 (PGP 9.5) and calcitonin gene-related peptide (CGRP). Nerves and vessels were counted and compared. Our results indicate that PGP 9.5(+) and CGRP(+) nerves were most numerous in growing hemangiomas and numbers were reduced in involuting hemangiomas and vascular malformations. The percentage of CGRP(+) sensory nerves was markedly increased in growing hemangiomas (45.3%) compared with involuting hemangiomas (21.2). These data indicate that hemangiomas with increasing neovascularization have increased sensory nerve growth. Sensory nerve-derived neuropeptides are known to act as endothelial cell mitogens and may contribute to the angiogenesis in these vascular tumors. Conversely, angiogenic endothelial cells may secrete mediators that promote nerve fiber growth. These results suggest that endothelial cell proliferation and sensory nerve fiber growth may be closely related.

Blood Vessels↗

PubMed2000

Vitronectin deficiency is associated with increased wound fibrinolysis and decreased microvascular angiogenesis in mice.

BACKGROUND: Vitronectin has several putative functions including regulating hemostasis, cell adhesion, and cell migration. However, the targeted deletion of vitronectin in mice results in normal development and normal coagulation parameters. To determine whether vitronectin may be necessary for nondevelopmental processes, we examined the response to tissue injury in vitronectin-null mice. METHODS: We examined wound healing in control and vitronectin-null mice by healing rate, zymography, reverse zymography, and Western blots. RESULTS: We found that dermal wound healing was slightly delayed in mice lacking vitronectin. More importantly, we found extensive areas of delayed hemorrhage near the sprouting tips of microvessels between days 7 and 14, which temporally coincided with increased urokinase-type plasminogen activator and tissue-type plasminogen activator activity by zymography. Though Western blots confirmed the presence of plasminogen activator inhibitor-1 protein throughout wound repair and reverse zymograms showed decreased plasminogen activator inhibitor-1 activity between days 7 and 14. CONCLUSIONS: Loss of vitronectin in mice was associated with changes in the fibrinolytic balance, and this may have led to focal sites of delayed hemorrhage. The mechanism that resulted in decreased angiogenesis and the formation of larger blood vessels in response to tissue injury remains unknown. This study suggests that vitronectin may have several distinct functions that are not required for normal development but are manifested in response to tissue injury.

Animals↗

PubMed2000

Current status of burn wound pathophysiology.

Healing is a continuum that can be unpredictable. Despite many advances and understanding of the multiple cellular processes and molecules involved in burn wound healing, physicians and patients have yet to reap the full benefit of this knowledge. The advances have occurred in a very short period, and with the exponential growth of molecular biology techniques and transgenic animal models, our understanding and treatment of burn wound healing could change exponentially over the next 10 years. The goal must be to continue to improve functional outcomes for burn survivors just as we have conquered critical care management for acutely injured burn patients.

Angiogenesis Inducing Agents↗

PubMed1999

Methemoglobinemia secondary to topical silver nitrate therapy--a case report.

Methemoglobinemia is a rare complication in individuals exposed to nitrates or nitrites. Whereas methemoglobinemia is a recognized potential complication in burn patients treated with topical 0.5% silver nitrate solution, no report of methemoglobinemia in burn patients has been present in the literature for more than 15 years. We raise consciousness about this complication with a case report of a 12-month-old child with necrotizing fasciitis resulting from a cutaneous flank infection. The patient developed cyanosis 20 days after initiation of topical treatment with 0.5% silver nitrate solution. Intravenous injection of methylene blue can restore normal blood oxygenation.

Administration, Topical↗

PubMed1999

Temporal activity of plasminogen activators and matrix metalloproteinases during cutaneous wound repair.

BACKGROUND: Response to tissue injury begins with the deposition of a fibrin-rich clot or the provisional matrix. The provisional matrix consists of plasma-borne matrix molecules that serve as scaffolding for the ensuing migration of cells. During wound repair multiple cell types must migrate through the clot-matrix scaffolding. The migration of these cells through the matrix is dependent on the activity of the fibrinolytic and proteolytic systems, which include the plasminogen activator (PA) system and matrix metalloproteinases (MMP). The aim of this study was to better understand the temporal activity of these enzymes during normal wound repair. METHODS: We used the murine excisional wound model and extracted proteins under nonreducing conditions. With use of gelatin and casein zymography, we determined the activity of the MMPs during the course of wound repair. In addition, we quantified the activity of MMP-2 and MMP-9 by a standardized assay. Plasminogen zymograms were used to detect urokinase PA and tissue PA activity. Western blots were used to detect the natural inhibitor of PAs, plasminogen activator inhibitor type 1. RESULTS: Our results demonstrate the temporal activity of MMP-2, MMP-3, MMP-7, and MMP-9 during the course of normal dermal repair. The activity of urokinase PA and tissue PA were also determined; it preceded the activity of the MMPs. CONCLUSIONS: We demonstrate the temporal activity of the 2 protease families, MMPs and PAs, in the normal process of cutaneous wound healing.

Animals↗

PubMed1998

Vitronectin decreases microvascular endothelial cell apoptosis.

Angiogenesis after tissue injury occurs in a matrix environment consisting of fibrin, fibronectin, and vitronectin as the major extracellular matrix (ECM) constituents. ECM-integrin interactions is critical for angiogenesis and failure to bind a ligand to certain integrin receptors (alpha[v]beta3 or alpha[v]beta5) inhibits angiogenesis. The ligand that binds to alpha(v)beta3 or alpha(v)beta5 integrin receptors during microvascular angiogenesis has not been identified. Our hypothesis is that provisional matrix molecules provide the environmental context cues to microvascular endothelial cells and promote angiogenesis by decreased programmed cell death. Using cultured human microvascular endothelial cells, we show that vitronectin, in comparison to growth on alternative provisional matrix molecules (fibronectin, fibrinogen plus thrombin), collagen I, and basement membrane molecules (collagen IV), significantly reduces microvascular endothelial cell death in vitro. This reduction was observed using morphologic criteria, TdT-mediated dUTP nick end labeling (TUNEL) assay, histone release into the cytoplasm, and thymidine release into the supernatant. Though our data confirm that vitronectin may bind to more than one integrin receptor to reduce MEC apoptosis, binding to the alpha(v) component appears to be the critical integrin subcomponent for reducing apoptosis.

Apoptosis↗

PubMed1998

Changes in matrix composition during the growth and regression of human hemangiomas.

BACKGROUND: Hemangiomas offer an uncommon opportunity to study rapid vessel growth and spontaneous regression of a vascular human tumor. In contrast, venous malformations are another type of vascular tumor that grows slowly without spontaneous involution. Extracellular matrix (ECM) molecules modulate the responsiveness of endothelial cells to mitogenic stimuli such as basic fibroblast growth factor (bFGF), a well-recognized stimulant of angiogenesis. In this study we hypothesized that in hemangiomas, sites of angiogenesis may have a different ECM composition than sites of vascular regression. MATERIALS AND METHODS: Using immunohistochemistry, we analyzed proliferating hemangiomas, regressing hemangiomas, venous malformations, and normal skin for the basement membrane ECM molecules collagen IV and laminin and plasma-borne ECM molecules fibronectin and vitronectin. We used metabolic labeling to determine whether primary human dermal microvascular endothelial cells regulated FGFR-1 or FGFR-2 when grown on these different matrices. RESULTS: We found that proliferating hemangiomas showed extensive deposition of vitronectin in the subendothelial space. In contrast, regressing hemangiomas or venous malformations did not show vitronectin deposition. Venous malformations, which are composed of ectatic lakes of venous channels, also lacked laminin in their basement membranes. We also found that cultured microvascular endothelial cells grown on vitronectin increased synthesis of FGFR-1 and FGFR-2 protein. CONCLUSIONS: Changes in the ECM environment occur in conjunction with the angiogenic state of a vascular human tumor. Furthermore, changes in the ECM environment alone can directly regulate synthesis of angiogenic growth factor receptors.

Collagen↗

PubMed1998

Protein gene product 9.5 is expressed by fibroblasts in human cutaneous wounds.

In a study initially designed to evaluate reinnervation of human cutaneous wounds using an antibody to the neuroneal marker protein gene product (PGP) 9.5, we observed marked immunostaining of cells with morphologic features of fibroblasts in the wounds. PGP 9.5 has recently been shown to be an important enzyme in the highly conserved ubiquitin system of proteolysis. Because the ubiquitin system is known to play an important role in regulating the cell cycle, the presence of PGP 9.5 in cells at a wound site was of considerable interest. Our objectives were to clarify the time frame for the appearance of PGP 9.5 and ubiquitin in wounds, to verify that PGP 9.5 is produced by wound fibroblasts, and to evaluate a potential role for these proteins in the tissue repair process. Standard incisional human wounds were stained with antibodies specific for PGP 9.5 and ubiquitin. At 7 d, stellate cells with morphologic features of fibroblasts stained for PGP 9.5, whereas earlier wounds were generally negative. In 14 and 21 d incised wounds and in chronic granulation tissue from nonhealing ulcers there was strong cellular staining for PGP 9.5 and for ubiquitin. These stellate cells also showed expression of mRNA for PGP 9.5 by reverse transcriptase-polymerase chain reaction in situ hybridization. PGP 9.5 was detected in cultured fibroblasts both by reverse transcriptase-polymerase chain reaction and by northern blot analysis. Confocal microscopy showed colocalization of antibodies to PGP 9.5 and prolyl-4-hydroxylase (a fibroblast marker) as well as colocalization of PGP 9.5 and the platelet derived growth factor beta receptor. We conclude that ubiquitin and PGP 9.5 were expressed by fibroblasts during the granulation tissue and remodeling phases wound healing. The mRNA for PGP 9.5 was demonstrated in stellate cells in chronic wounds and in fibroblasts in culture. The appearance of these degradative proteins in later wounds suggests a downregulation function in the wound healing response.

Apoptosis↗

PubMed1997

Monocyte chemoattractant protein-1 mRNA expression in the human burn wound.

The inflammatory response following a thermal insult begins with the skin itself. Langerhan's cells, tissue macrophages, keratinocytes, fibroblasts, and endothelial cells contribute to the initial events of wound healing with active and passive release of cell mediators. One of the mediators potentially important to the repair process is monocyte chemoattractant protein-1 (MCP-1). Macrophages, fibroblasts, endothelial cells, and keratinocytes can produce MCP-1 in response to inflammatory stimuli. Therefore, we evaluated 10 human burn wound specimens for MCP-1 mRNA using in situ hybridization. Selected specimens of different ages were examined using combined in situ hybridization and immunocytochemistry to identify cell types that expressed MCP-1 mRNA. Antibodies to HAM56 for macrophages, CD45 for bone marrow-derived cells, Factor VIII for endothelial cells, and Factor XIIIa for dermal antigen-presenting cells were included in these experiments. By Postburn Day 2, basal layer keratinocytes at the edges of the wound had upregulated MCP-1 message; the increased signal persisted in the rate pegs deep in the dermal wound bed through 49 days postinjury. Occasional FXIIIa+ immunostained dermal cells expressed MCP-1 mRNA. Islands of granulation tissue throughout the wound bed were positive for increased expression of MCP-1; endothelial cells and inflammatory cells both contributed to this upregulated signal. Our data support the theory that the skin itself is a component of the immune system and that noninflammatory cells contribute to the initiation and maintenance of the inflammation at a wound site. Failure to produce MCP-1 or other related mediators by indigenous cutaneous cells may delay the inflammatory response to injury and potentially disrupt other essential phases of wound repair.

Antibodies↗

PubMed1996

Ontogeny and characterization of factor XIIIa+ cells in developing human skin.

Factor XIIIa (FXIIIa), a coagulation transglutaminase, is a cytoplasmic marker for dermal dendritic cells reported to be bone marrow-derived, phagocytic and antigen-presenting. In non-inflamed skin, these cells populate the papillary dermis in a perivascular distribution. They are increased in dermatoproliferative disorders and have been implicated as dermal stimulants for psoriatic hyperkeratosis. Since developing skin provides an example of dermal influence on the epidermis, we evaluated the presence of FXIIIa+ cells in human fetal skin to determine whether their location would suggest a role in morphogenetic events in the skin. Embryonic and fetal skin of progressive estimated gestational ages (EGA) was examined using immunocytochemistry with a polyclonal antibody to FXIIIa. At 6 weeks EGA, globular FXIIa+ cells were present in the hypodermis. By 7-8 weeks, a compact sub-epidermal network of fusiform FXIIIa+ cells was also evident. By 11-12 weeks, the sub-epidermal cellular network was no longer FXIIIa+, but discrete FXIIIa+ dendritic cells were present in the reticular dermis. With advancing gestational age, FXIIIa+ dendritic cells populated the papillary dermis in a perivascular distribution. This adult-like distribution persisted through 22 weeks EGA, the oldest specimen examined. Because FXIIIa+ cells were evident in embryonic skin before the onset of bone marrow hematopoietic function, the skin was double-labeled with the FXIIIa antibody and with monoclonal antibodies to CD45 (marker for bone marrow-derived cells), CD68 (marker for macrophages) and HLA-DR (class II major histocompatibility antigen). Most of the FXIIIa+ dendritic cells did not colocalize CD45, but were CD68+; some cells did react with the HLA-DR antibody. Notably, the FXIIIa+ cells of the sub-epidermal network in the 7 weeks EGA specimens did not react with the other antibodies. We conclude that FXIIIa+ cells are first present in embryonic hypodermis and sub-epidermal dermis and later they are distributed in the papillary dermis in a perivascular pattern. In embryonic skin FXIIIa+ cells are not exclusively dendritic. Our data support the idea that cells that express FXIIIa do not constitute a unique bone marrow-derived cell type, but that multiple cell types produce FXIIIa.

Antibodies, Monoclonal↗

PubMed1995

Immunolocalization of FXIIIa+ dendritic cells in human burn wounds.

Dermal dendrocytes constitute a population of indigenous antigen-presenting cells that have been implicated in dermal inflammation and may have a role in wound healing. They are identified by expression of the transglutaminase coagulation Factor XIIIa (FXIIIa) and a perivascular distribution in the papillary dermis. In this study, we used immunohistochemistry to localize and characterize FXIIIa+ cells in healing burn wounds. Wound specimens from Postburn Days 2, 3, 4, 5, 6, 8, 9, 10, 11, 15, 30, and 49 were collected from 18 patients at the time of excision and grafting, processed for immunolabeling, and labeled with an antibody to FXIIIa. Tissue sections were also double-labeled with the anti-FXIIIa and with antibodies to CD68, specific for macrophages, CD45, specific for bone marrow-derived cells, and proliferating cell nuclear antigen (PCNA) for proliferation. Antigen-presenting status was evaluated using an antibody to the major histocompatibility complex HLA-DR. The dermis subjacent to the tongue of proliferating epithelium at the wound edge had increased numbers of FXIIIa+ dendritic cells compared to the cellular distribution in uninjured skin. FXIIIa+ dendritic cells were absent from the burned dermis on all postburn days examined in this study. However, capillaries in the deep dermis had a FXIIIa+ granular staining pattern. CD68+ cells and CD45+ cells were present throughout the wound bed at all stages of healing, indicating an inflammatory cell response in the injured dermis. Antibodies to PCNA did not colocalize to FXIIIa+ cells, suggesting that the dermal dendrocytes were not proliferating. The antibody to HLA-DR localized to some, but not all of the FXIIIa+ dendritic cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Biomarkers↗

PubMed1994

Basic fibroblast growth factor in the early human burn wound.

The role of endogenous growth factors in normal wound healing is not clear. Most of the data on growth factors in healing wounds have been obtained from the application of recombinant exogenous growth factors to animal and human wounds. We describe the immunolocalization of basic fibroblast growth factor (bFGF) in the injured dermis of skin from patients with partial and full-thickness burns. Three antibodies demonstrate an extracellular staining pattern of bFGF corresponding to areas of tissue injury that was most intense in specimens collected between 4 and 11 days post-burn injury. In contrast, bFGF staining appeared markedly decreased by Postburn Day 17 and was more consistent with uninjured tissue in a 30-day-old burn that had virtually reepithelialized. Basic FGF staining in the non-burned skin from the same patients was restricted to the dermal capillary basement membranes and the sweat glands, which is consistent with other reports of immunoreactive bFGF localization in normal adult skin. The immunohistochemical results were confirmed with Western immunoblots of the same tissue. The major band at 16.5 kDa, which is within the recognized range of the bFGF molecule's several forms, was detected in both burned and unburned tissue from the same patient. These findings support the hypothesis that bFGF is a presynthesized mediator that is stored in either the cells or extracellular matrix, is released locally from sites of direct injury, and may be important in early wound healing.

Blotting, Western↗

PubMed1994

Engine block burns: Dupuytren's fourth-, fifth-, and sixth-degree burns.

We recently treated two patients with engine block-muffler contact burns and greatly underestimated the devastating injuries to bone, deep fascia, and muscle. As a result, each patient required multiple procedures to close their burn wounds. Ten-year data from the University of Washington Burn Unit confirmed our observation that these burns tend to be considerably deeper than suspected. Eighteen patients with contact burns from engine parts were identified from 1980 through 1990. Nine (50%) of these were initially recognized to be fourth-degree and five (28%) were third-degree thermal injuries, showing that these are deep burns. Eight patients required fascial excisions and four required debridement of devascularized bone. The mean burn size was only 6% total body surface area; however, the patients with fourth-degree burns had an average graft take of only 56% and required a mean hospital stay of 44 days. Patients with third-degree burns also had suboptimal graft take and some required prolonged hospitalization. Thirty-six percent of patients required flaps either as the initial procedure or as a second procedure following an autograft. The four patients with partial-thickness burns healed without surgery and their average length of hospital stay was 3 days. Of the entire group, only four healed without surgery and only five healed with a single operation. Our 10-year data indicate that engine block contact burns are usually small, but most are deceptively deep, involving tendon, muscle, or bone. If the burn appears full thickness, suspicion must be very high at the initial surgical procedure that there is deep tissue destruction.(ABSTRACT TRUNCATED AT 250 WORDS)

Accidents, Traffic↗

PubMed1993

Mediators in thermal injury.

The combined goal of each of the mediators discussed here is homeostasis in a stressful situation. The associated immunosuppression and immunohyperactivity are unfortunate sequelae of massive upregulation of mediators normally produced in miniscule quantities. Perhaps they were once necessary to regulate survival of severely injured patients. However, Stoner's question about whether these mediators are beneficial for modern medical practices is very relevant. Our significant challenge is now to counteract the body's own efforts to heal itself. Understanding the interactions initiates this task; discovering therapeutic control mechanisms must be the next step.

Arachidonic Acid↗

PubMed1992

Iliofemoral venous thrombosis following fascial excision of a deep burn of the lower extremity: case report.

Burned patients with deep venous thrombosis present a particularly perplexing challenge. They frequently require central venous catheters. Their altered skin integrity does not permit correlation with the typical changes described by the classic terminology for thrombophlebitis or its most severe forms, phlegmasia cerulea dolens or phlegmasia alba dolens. They are at risk of exsanguination or massive graft loss with lytic therapy or anticoagulation. Venous thrombectomy may be a necessary limb-saving surgical option.

Adult↗

PubMed1991

Novel approach to iatrogenic bile peritonitis.

Bile peritonitis after injury to the biliary tree is a serious complication that requires exploratory laparotomy. Our patient had an obstructing ampullary carcinoma, and generalized bile peritonitis developed from attempted percutaneous transhepatic cholangiography. The patient's condition was managed by peritoneal lavage and endoscopic transampullary stenting, with immediate relief of pain and toxicity. Exploratory laparotomy was avoided, and an eventual pylorus-sparing Whipple resection was the definitive treatment. We believe this to be the first report of successful nonoperative treatment of a patient with bile peritonitis with obstructive jaundice.

Adenocarcinoma↗

PubMed1982

Characterization of a postlavage, in situ pulmonary macrophage population.

A postlavage in situ subpopulation of pulmonary macrophages (PM), biochemically distinct from the lavaged population, has recently been isolated from rats. After exhaustive bronchopulmonary lavage to extract the free lung cells, the lungs were excised, homogenized, and filtered, and the resultant cell suspension was allowed to form a monolayer on plastic Petri dishes. Electron microscopic morphometry failed to indicate any morphologic differences in the two populations. The postlavage in situ PM were more active metabolically during phagocytosis of zymosan particles or stimulation by phorbol myristate acetate (PMA) than the corresponding lavage population, as evidenced by greater superoxide generation. Macrophages prepared by either method became more avidly phagocytic when incubated with cell-free medium isolated in the preparation of the situ population. Peroxidase, an enzyme absent from the granules of PM separated by lavage techniques, was found in a granule-rich fraction of the in situ macrophage. Catalase activity was found in similar amounts in both supernatants and granule-rich fractions of both populations. The results support the concept of subpopulations of PM and suggest that these subpopulations are distinguished by their biochemical properties and their functional abilities.

Animals↗