Outcomes of unrelated cord blood transplants and allogeneic-related hematopoietic stem cell transplants in children with high-risk acute lymphocytic leukemia.

Acute lymphocytic leukemia (ALL) is a common indication for hematopoietic stem cell transplantation (HSCT) in children. Use of unrelated cord blood (UCB) has become increasingly popular as a stem cell source, given the rapid availability and decreased GVHD potential. Publications describing outcomes of children with leukemia who underwent UCB transplants have compared them to those having received unrelated donor marrow transplants. Results are similar. We compared our outcomes using UCB vs allogeneic-related hematopoietic stem cells in pediatric ALL patients since 1992. A total of 49 patients were analyzed. All patients were either in CR1 with high-risk features (n=21) or in CR2 (n=28) with initial remission less than 36 months. Patients received myeloablation with fractionated total body irradiation, cyclophosphamide, and etoposide and GVHD prophylaxis with cyclosporine and methotrexate. Antithymocyte globulin was added for UCB recipients to address the HLA differences. In all, 23 patients underwent allogeneic -related HSCT and 26 underwent UCB transplantation. Other than increased time to engraftment for UCB recipients, results are equivalent. The 3-year overall survival is 64% and 3-year event-free survival is 60% for both groups. Rates of GVHD and transplant-related mortality are also equivalent. UCB is a reasonable option for children with ALL who are referred for HSCT.

Severe idiopathic gastroparesis due to neuronal and interstitial cells of Cajal degeneration: pathological findings and management.

Delayed gastric emptying can be due to muscular, neural, or humoral abnormalities. In the absence of an identified cause, gastroparesis is labelled as idiopathic. We present the case of a patient with severe idiopathic gastroparesis. Pharmacological approaches failed, as well as reduction in gastric emptying resistance with pyloric injection of botulinum toxin and pyloroplasty. Therefore, subtotal gastrectomy was performed. Histological and immunohistochemical study of the resected specimen showed hypoganglionosis, neuronal dysplasia, and a marked reduction in both myenteric and intramuscular interstitial cells of Cajal. To our knowledge, this is the first time these rare histological findings have been described in a patient with idiopathic gastroparesis.

Accumulation of mast cells and macrophages in focal active gastritis of patients with Crohn's disease.

BACKGROUND/AIMS: Recent studies have shown that focal active gastritis seems to be the typical gastric pathology in Crohn's disease. The aim of this study was to compare the incidence of focal active gastritis, Helicobacter pylori infection and distribution of gastric mast cells and macrophages in patients with Crohn's disease, ulcerative colitis and H. pylori gastritis without inflammatory bowel disease. METHODOLOGY: Patients with histologically confirmed Crohn's disease (n = 25) or ulcerative colitis (n = 25) and control patients without inflammatory bowel disease (n = 25) were included in this study. Biopsy specimens were obtained from the antrum and corpus of each patient, and stained with hematoxylin and eosin and immunostained using antibodies to tryptase (AA1) and CD68. The number of mast cells and macrophages located in the lamina propria was determined. RESULTS: Focal active gastritis was detected in 54% of H. pylori-negative patients with Crohn's disease, but it was not found in patients with ulcerative colitis nor in the control group. The density of mast cells and macrophages in the lamina propria of H. pylori-positive patients was significantly higher than in H. pylori-negative patients in all groups. In the Crohn's disease group, the number of mast cells (antrum; 83 +/- 11, body; 89 +/- 11/mm2) and macrophages (antrum; 94 +/- 22, body; 92 +/- 17/mm2) in the lamina propria of H. pylori-negative patients with focal active gastritis was halfway between that in H. pylori-positive and H. pylori-negative patients. In focal active gastritis, mast cells accumulated at the border of focal active gastritis, whereas macrophages accumulated in the center of such lesions. CONCLUSIONS: Our findings indicated that the diagnosis of focal active gastritis, using immunostain for mast cells and macrophages, is the histological hallmark of gastric Crohn's disease. Macrophages might be associated with the formation of focal active gastritis in patients with Crohn's disease.

Gastrointestinal stromal tumors may originate from a subset of CD34-positive interstitial cells of Cajal.

Most gastrointestinal stromal tumors (GISTs), a subgroup of mesenchymal neoplasms of the gut wall, express both Kit (CD117) and CD34 proteins. It has been suggested that GISTs originate from or differentiate into interstitial cells of Cajal (ICC), after several reports indicated that ICC are likely the only cells in the gut which express both Kit and CD34. ICC are among the few cell types resident in the gut which express Kit, together with mast cells. However, the question whether or not ICC express CD34 is currently disputed. Using single-cell reverse transcriptase-polymerase chain reaction (RT-PCR) on cultured murine intestinal cells, single ICC were selected by morphology and tested for the expression of c-kit and CD34 mRNA. Most ICC were only c-kit-positive, however a subset (7 out of 43) were double positive for both c-kit and CD34. In the human small intestine, sequential immunohistochemical staining for Kit and CD34 proteins on the same 3-microm sections showed that some of the ICC surrounding Auerbach's plexus and ICC within the circular muscle layer of the small intestine were positive for both Kit and CD34. In addition, CD34(+)Kit(-) cells were seen adjacent to ICC. These data from two different techniques indicate that ICC can be double positive for Kit and CD34. Thus, GISTs with the Kit(+)CD34(+) phenotype may arise from a subpopulation of CD34(+) Kit(+) ICC.

Transgenic expression of granulocyte-macrophage colony-stimulating factor induces the differentiation and activation of a novel dendritic cell population in the lung.

The role of granulocyte-macrophage colony-stimulating factor (GM-CSF) in the differentiation of dendritic cells (DCs) during pulmonary viral infection was investigated by using a mouse model of GM-CSF transgene expression established with an adenoviral vector (AdGM-CSF). GM-CSF gene transfer resulted in increased levels of GM-CSF in the lung, which peaked at day 4 and remained increased up to day 19. A striking cellular response composed predominantly of macrophage-like cells was observed in the lung receiving AdGM-CSF but not control vector. By FACS analysis, the majority of these cells were identified at an early time point as macrophages and later as mature/activated myeloid DCs characterized by CD11b(bright), CD11c(bright), MHC class II(bright), and B7.1(bright). In contrast, GM-CSF had a weak effect on a small DC population that was found present in normal lung and was characterized by CD11c(bright) and CD11b(low). By immunohistochemistry staining for MHC II, the majority of activated antigen-presenting cells were localized to the airway epithelium and peribronchial/perivascular areas in the lung. A concurrently enhanced Th1 immune response was observed under these conditions. The number of CD4 and CD8 T cells was markedly increased in the lung expressing GM-CSF, accompanied by increased release of interferon (IFN)gamma in the lung. Furthermore, lymphocytes isolated from either lung parenchyma or local draining lymph nodes of these mice but not the control mice released large amounts of IFNgamma on adenoviral antigen stimulation in vitro. These findings reveal that GM-CSF promotes the differentiation and activation of a myeloid DC population primarily by acting on macrophages during pulmonary immune responses.

Immunohistochemical evidence for estrogen receptors in meibomian glands.

PURPOSE: To look for sex hormone receptor distribution in three structures contributing to the normal human tear film: the conjunctiva, the accessory lacrimal glands, and the meibomian glands. DESIGN: An immunohistochemical study. TISSUES AND CONTROLS: Forty-one upper eyelid specimens were collected from 15 male and 26 female patients (age range, 1.5-85 years) during blepharoptosis surgery via posterior tarsoconjunctival mullerectomy (Fasanella-Servat or Gavaris). In addition, control sections of histologically normal breast, prostate, and skin tissue were obtained. METHODS: Immunohistochemical staining using mouse monoclonal antibodies against estrogen, progesterone, and androgen receptors was performed on all tissues and controls. Quantitation of the receptors was performed and expressed as percentage nuclear positivity. Specimens were divided into three groups based on the age of the patient: <12 years (n = 9); 18-55 years (n = 1); >55 years (n = 12). RESULTS: Forty-one specimens contained conjunctiva. All were negative for estrogen, progesterone, and androgen receptors. Twenty-four specimens contained accessory lacrimal glands of Wolfring. All were negative for the three receptors. Twenty-two specimens contained meibomian glands. All were positive for estrogen receptors; one was positive for progesterone receptors and one for androgen receptors. Using Minitab statistical software (Minitab Inc. State College, PA), analysis of variation revealed no statistical difference between sexes or between age groups studied. The sebaceous glands of skin were uniformly positive for androgen receptors. Sebaceous glands of the face and scalp (3 of the 15 skin samples) were also positive for estrogen receptors. CONCLUSIONS: Estrogen receptors are present in the meibomian glands of the upper eyelid. Unlike sebaceous glands elsewhere on the skin, the meibomian glands lack androgen receptors. Estrogen receptors may play a role in modulation of the lipid layer of the tear film, and their activity may be linked to meibomian gland dysfunction and dry eye syndrome.

Characterization of adrenergic receptors in the accessory lacrimal glands of the upper eyelid.

PURPOSE: To identify the histologic location as well as the exact subtype of adrenergic receptors in the accessory lacrimal glands of the upper eyelid. METHODS: Upper eyelid specimens were collected from 19 patients undergoing routine blepharoptosis correction via a posterior tarsoconjunctival mullerectomy. Immunohistochemical staining using polyclonal antibodies against human alpha 1, alpha 2, beta 1, and beta 2 receptors was performed on all specimens. RESULTS: beta 1 receptors were the predominant adrenergic receptor subtype in the glands of Wolfring. CONCLUSIONS: The presence of beta 1 receptors in the accessory lacrimal glands of the upper eyelid may suggest a possible role for selective beta 1 agonists in the treatment of keratitis sicca.

Distribution of adrenergic receptor subtypes in the retractor muscles of the upper eyelid.

PURPOSE: To identify adrenergic receptor subtypes and their relative distribution in the retractor muscles of the upper eyelid, the levator palpebrae superioris, and the Müller muscle. The pattern of distribution of these receptors in the Müller muscle was further compared in patients with dysthyroid eyelid retraction and in normal subjects. METHODS: Müller muscle specimens were collected from 19 patients undergoing ptosis correction and from 8 patients undergoing repair of dysthyroid eyelid retraction. Immunohistochemical staining for alpha 1-, alpha 2-, beta 1-, and beta 2-adrenergic receptors was performed using antihuman rabbit polyclonal antibodies. RESULTS: alpha 2-Adrenergic receptors were the predominant subtype in the Müller muscle, and beta 1-adrenergic receptors were the predominant subtype in the levator muscle. There was no significant difference in the staining pattern between specimens collected from patients with dysthyroid eyelid retraction and those from normal subjects. CONCLUSIONS: The interaction between the alpha 2 and beta 1 receptors in the upper eyelid retractor muscles may be important in the control of the upper eyelid position and may contribute to the development of dysthyroid eyelid retraction. Specific alpha 2 antagonists could be developed and may be effective pharmacologic agents for the treatment of eyelid retraction.

Interstitial cells of Cajal as precursors of gastrointestinal stromal tumors.

Interstitial cells of Cajal (ICC) are implicated in the regulation of gut peristalsis and are immunostained by antibodies against Kit (CD117), a tyrosine kinase receptor. Most gastrointestinal mesenchymal tumors (GIMTs) are of uncertain histogenesis, although many are CD34-positive. CD34 was found to colocalize with vimentin (Vim) and the Kit-positive networks of cells within and around neural plexi, indicating that ICC can be Vim- and CD34-positive. ICCs appear to be the only Kit+CD34+Vim+ cell in the gut. Formalin-fixed, paraffin-embedded tissues from 43 GIMTs were immunostained for Kit, CD34, Vim, PGP 9.5 (PGP, a neural marker), muscle-specific actin (MSA), and other markers including desmin (Des). Eight tumors were myoid (MSA+Des+Vim-Kit-CD34-), and one was a schwannoma (PGP+S100+Vim+Kit-CD34-), but 34 tumors were of uncertain histogenesis (gastrointestinal stromal tumors, GIST), exhibiting neither a complete myoid nor a schwannian immunophenotype. All 34 were Vim+, and 33/34 were either Kit (n = 30) or CD34 (n = 23) immunoreactive. Of these 34 GIST, 24 were negative for all myoid and neural markers, 6 were PGP+S100-, and 4 were MSA+Des-. The Kit+CD34+Vim+ immunophenotype of GIST suggests that they originate from, or have differentiated into, ICC-like cells; the term ICC tumor (ICCT) is suggested. Kit is a more sensitive marker than CD34 for ICCT, but both are required in tumor identification. All clinically malignant GISTs were pathologically malignant (size, mitoses) but also showed loss of either CD34 or Kit. "Blind" examination of electron micrographs in 10 tumors showed them to be heterogeneous. Some had features seen in normal ICC, but cells could not be positively identified as being adult ICC. GIMT may therefore be classifiable into those with pure myoid, schwannian (or neural) differentiation, but the majority are of ICC origin or show ICC differentiation immunophenotypically (ICCT).

Transfer of tumor necrosis factor-alpha to rat lung induces severe pulmonary inflammation and patchy interstitial fibrogenesis with induction of transforming growth factor-beta1 and myofibroblasts.

Tumor necrosis factor-alpha is up-regulated in a variety of different human immune-inflammatory and fibrotic pulmonary pathologies. However, its precise role in these pathologies and, in particular, the mechanism(s) by which it may induce fibrogenesis are not yet elucidated. Using a replication-deficient adenovirus to transfer the cDNA of tumor necrosis factor-alpha to rat lung, we have been able to study the effect of transient but prolonged (7 to 10 days) overexpression of tumor necrosis factor-alpha in normal adult pulmonary tissue. We have demonstrated that local overexpression resulted in severe pulmonary inflammation with significant increases in neutrophils, macrophages, and lymphocytes and, to a lesser extent, eosinophils, with a peak at day 7. By day 14, the inflammatory cell accumulation had declined, and fibrogenesis became evident, with fibroblast accumulation and deposition of extracellular matrix proteins. Fibrotic changes were patchy but persisted to beyond day 64. To elucidate the mechanism underlying this fibrogenesis, we examined bronchoalveolar fluids for the presence of the fibrogenic cytokine transforming growth factor-beta1 and tissues for induction of alpha-smooth muscle actin-rich myofibroblasts. Transforming growth factor-beta1 was transiently elevated from day 7 (peak at day 14) immediately preceding the onset of fibrogenesis. Furthermore, there was a striking accumulation of myofibroblasts from day 7, with the most extensive and intense immunostaining at day 14, ie, coincident with the up-regulation of transforming growth factor-beta1 and onset of fibrogenesis. Thus, we have provided a model of tumor necrosis factor-alpha-mediated pulmonary inflammation and fibrosis in normal adult lung, and we suggest that the fibrogenesis may be mediated by the secondary up-regulation of transforming growth factor-beta1 and induction of pulmonary myofibroblasts.

Cytokeratin 18 is expressed on the hepatocyte plasma membrane surface and interacts with thrombin-antithrombin complexes.

During experiments to identify putative hepatic receptors for thrombin-antithrombin (TAT) complexes, a 45-kDa protein was identified by ligand blotting. Following gel purification, amino acid sequencing revealed the 45-kDa TAT-binding polypeptide to be cytokeratin 18 (CK18). The presence of CK18 on the surface of intact rat hepatoma cells was demonstrated by binding of 125I-anti-CK18 antibodies. Anti-CK18 antibodies reduced the binding and internalization of 125I-TAT by rat hepatoma cells. Immunocytochemical analysis, to determine the location of CK18 in vivo, revealed a periportal gradient of CK18 staining; with hepatocytes around the portal triads demonstrating striking pericellular staining. In addition, anti-CK18 IgG associated with perfused livers to a significantly greater extent than preimmune IgG. Taken together, these data provide evidence that CK18 is found on the extracellular surface of hepatocytes and could play a role in TAT removal. Finally, these data, in conjunction with recent reports of CK8 (Hembrough, T. A., Li, L., and Gonias, S. L. (1996) J. Biol. Chem. 271, 25684-25691) and CK1 cell membrane surface expression (Schmaier, A. H. (1997) Thromb. Hemostasis 78, 101-107), indicate a novel role for these proteins as putative cellular receptors or cofactors to cellular receptors.

Mast cell Fc epsilonRI expression in the rat intestinal mucosa and tongue is enhanced during Nippostrongylus brasiliensis infection and can be up-regulated by in vivo administration of IgE.

The activation of rodent and human mast cells can occur through the cross-linking of tetrameric IgE receptors each containing single alpha- and beta- and two gamma-subunits. However, the factors that regulate the in vivo expression of Fc epsilonRI are poorly understood. We have examined the expression of the Fc epsilonRI beta-subunit in the Nippostrongylus brasiliensis (Nb)-induced mode l of rat intestinal inflammation. We developed a double-staining technique for mast cell granules (Alcian blue) and the beta-subunit of Fc epsilonRI. The intensity of immunohistochemical staining per mast cell was quantified using an image analysis system. Jejunal and tongue mast cells of Lewis rats were visible by Alcian blue staining before Nb infection, but they expressed very low levels of beta-subunit as assessed by immunohistochemical staining. These levels were increased by day 11 postinfection and reached a maximum at day 14. Since serum IgE levels correlated well with the degree of beta-subunit expression, we investigated whether the observed enhancement of receptor expression might occur through the stabilization of receptor complexes by IgE. Therefore, Lewis rats were treated with myeloma IgE, and beta-subunit expression was examined. In both tongue and jejunal tissue, a significant rise in beta-subunit expression was observed in response to IgE injection, although levels of beta-subunit expression were not as high as those observed in Nb-infected animals. The increase in beta-subunit expression was accompanied by an increase in the amount of mast cell-associated IgE. These observations may have important implications for the regulation of IgE receptor expression during disease.

Intradermal transgenic expression of granulocyte-macrophage colony-stimulating factor induces neutrophilia, epidermal hyperplasia, Langerhans' cell/macrophage accumulation, and dermal fibrosis.

Granulocyte-macrophage colony-stimulating factor (GM-CSF), a pleiotropic cytokine, is up-regulated in a number of chronic skin inflammatory diseases, particularly atopic dermatitis. However, its role in these conditions remains largely unclear. To explore its function, we have established a rat intradermal transgene model by using a replication-deficient adenoviral vector expressing GM-CSF. Intradermal GM-CSF gene transfer led to a prolonged compartmentalized expression of transgene protein in the dermis. This expression induced an unexpectedly wide spectrum of pathologies in both epidermis and dermis, including neutrophilia, epidermal hyperplasia (acanthosis), an increased number of epidermal Langerhans' cells, accumulation of MHC II-positive macrophages, as well as mild eosinophilia in the dermis at earlier stages and upper dermal fibrosis at later stages. These findings thus identify GM-CSF as a potent multifunctional cytokine at skin site that is capable of evolving numerous inflammatory processes ranging from the early acute neutrophilia to later chronic fibrotic responses, and also suggest the important role of this cytokine in the development and perpetuation of pathologic changes in chronic skin inflammatory conditions including chronic atopic dermatitis. In addition, our study presents a novel model of adult normal animals that is useful for identifying and studying key cytokines involved in inflammatory skin diseases.

Community-based approach to schistosomiasis control.

With few exceptions, efforts to control schistosomiasis have relied upon ongoing community cooperation with "outsiders' rather than creating within the community the capacity and means for carrying out ongoing disease control measures with minimal external support. Offered as a useful model is a program in Kaele subdivision, Extreme North Province, Cameroon designed to establish and integrate within the primary health care (PHC) system the control of urinary schistosomiasis, hyperendemic in the region. At the community level, and with minimal dependence upon external resources, culturally appropriate and effective health education was instituted, the capacity to diagnose and treat schistosomiasis was created, diagnosis and drug therapy (praziquantel) was made available conveniently and at low cost, and, on a very limited basis, snails were controlled. Efforts were made to build upon and strengthen existing community structures and institutions rather than create new ones. The impact of the interventions was measured in terms of changes in knowledge and behavior, prevalence and intensity of infection, utilization of health services, and the ability to finance the control activities within the context of a generalized cost recovery system. Program successes and failures are discussed, as well as lessons learned and their implications.