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Biomedical subjects

Laura P Hale

Publications and source records attributed to Laura P Hale.

At least 19 recordsLinked to original sources
PubMed2006

Oral immunogenicity of the plant proteinase bromelain.

Bromelain is a natural mixture of proteolytic enzymes derived from pineapple stem that has been shown to have anti-inflammatory activity when administered orally. Although most proteins given orally without adjuvant (e.g., food) result in tolerance, we previously reported that long-term oral exposure to bromelain stimulated the development of high serum anti-bromelain antibody titers. The purpose of these studies was to further investigate the mechanisms responsible for the immunogenicity of oral bromelain. Results showed that repeated exposure was required for development of anti-bromelain antibodies, with strong antibody responses in all mice that received at least 12 doses of bromelain either orally or intragastrically over 3-6 weeks. Proteolytic activity was required for strong oral immunogenicity in the absence of conventional adjuvant, with strong serum antibody responses generated against proteolytically active bromelain and trypsin, but not against ovalbumin, lysozyme, or inactivated bromelain. Significantly higher anti-bromelain antibody titers were seen in IL-10-deficient versus wild-type mice, suggesting that simultaneous treatments that decrease IL-10 activity may further enhance systemic antibody responses following oral exposure. The antibodies generated did not affect the proteolytic activity of bromelain. The data demonstrate that proteolytically active antigens such as bromelain can stimulate both systemic and mucosal immune responses following repeated oral exposure. Further studies of the mechanisms involved in generation of immune responses following oral exposure to proteolytically active antigens can lead to a better understanding of mechanisms of oral tolerance and to the development of novel adjuvants for oral vaccines.

Adjuvants, Immunologic↗
PubMed2006

Neuronal expression of the Ccm2 gene in a new mouse model of cerebral cavernous malformations.

Cerebral cavernous malformations are vascular defects of the central nervous system consisting of clusters of dilated vessels that are subject to frequent hemorrhaging. The genes mutated in three forms of autosomal dominant cerebral cavernous malformations have been cloned, but it remains unclear which cell type is ultimately responsible for the lesion. In this article we describe mice with a gene trap insertion in the Ccm2 gene. Consistent with the human phenotype, heterozygous animals develop cerebral vascular malformations, although penetrance is low. Beta-galactosidase activity in heterozygous brain and in situ hybridization in wild-type brain revealed Ccm2 expression in neurons and choroid plexus but not in vascular endothelium of small vessels in the brain. The expression pattern of Ccm2 is similar to that of the Ccm1 gene and its interacting protein ICAP1 (Itgb1bp1). These data suggest that cerebral cavernous malformations arise as a result of defects in the neural parenchyma surrounding the vascular endothelial cells in the brain.

Animals↗
PubMed2005

Naïve T cells are maintained in the periphery during the first 3 months of acute HIV-1 infection: implications for analysis of thymus function.

A key determinant of T cell dynamics in HIV-1 infection is the status of thymic function. To date, most studies of the impact of HIV-1 on the thymus during early HIV-1 infection have been done in samples collected in the interval of 3-12 months after infection. In this study, we have probed the status of thymic function and peripheral naive T cells in patients with acute HIV-1 infection diagnosed 18-72 days after the onset of symptoms. We found that peripheral CD4 and CD8 T cell proliferation was initially elevated, then waned over time. The fall in T cell proliferation correlated with a reduction in HIV-1 viral RNA levels and a rise in peripheral blood CD4+ CD25+ T cells. In spite of elevated T cell proliferation early on in primary HIV-1 infection, levels of naive phenotype CD4 and CD8 T cells and T cell receptor excision circle positive cells (sjTREC(+)) remained constant. Taken together with the observation that T cell proliferation normally dilutes peripheral T cell episomal sjTREC levels, these data suggested that thymopoiesis contributes to maintenance of the naive T cell pool during the earliest stages of HIV-1 infection (18-72 days).

Adult↗
PubMed2005

Treatment with oral bromelain decreases colonic inflammation in the IL-10-deficient murine model of inflammatory bowel disease.

Bromelain is a mixture of proteinases derived from pineapple stem that is marketed in health food stores as a "digestive aid". Orally administered bromelain was anecdotally reported to induce clinical and endoscopic remission of ulcerative colitis in two patients whose disease was refractory to multi-agent conventional medical therapy. However, the potential efficacy of bromelain in colitis has not yet been tested rigorously in either animals or humans. In this study, the clinical and histologic severity of inflammatory bowel disease (IBD) was determined in IL-10-/- mice treated orally with bromelain in vivo. Daily treatment with oral bromelain beginning at age 5 weeks decreased the incidence and severity of spontaneous colitis in C57BL/6 IL-10-/- mice. Bromelain also significantly decreased the clinical and histologic severity of colonic inflammation when administered to piroxicam-exposed IL-10-/- mice with established colitis. Proteolytically active bromelain was required for anti-inflammatory effects in vivo. Adverse effects of dermatitis, hair loss, and weight loss due to mucositis were rare, dose related, and were not seen in wild-type mice treated orally with up to 1000 mg bromelain/kg/day for 18 weeks. Although the exact mechanisms by which exogenous proteinases affect bowel inflammation have not yet been determined, the results justify additional studies of this complementary biologically based approach to treatment of IBD.

Administration, Oral↗
PubMed2005

Proteinase activity and stability of natural bromelain preparations.

Bromelain is a complex mixture of proteinases typically derived from pineapple stem. Similar proteinases are also present in pineapple fruit. Beneficial therapeutic effects of bromelain have been suggested or proven in several human inflammatory diseases and animal models of inflammation, including arthritis and inflammatory bowel disease. However, it is not clear how each of the proteinases within bromelain contributes to its anti-inflammatory effects in vivo. Previous in vivo studies using bromelain have been limited by the lack of assays to control for potential differences in the composition and proteolytic activity of this naturally derived proteinase mixture. In this study, we present model substrate assays and assays for cleavage of bromelain-sensitive cell surface molecules can be used to assess the activity of constituent proteinases within bromelain without the need for biochemical separation of individual components. Commercially available chemical and nutraceutical preparations of bromelain contain predominately stem bromelain. In contrast, the proteinase activity of pineapple fruit reflects its composition of fruit bromelain>ananain approximately stem bromelain. Concentrated bromelain solutions (>50 mg/ml) are more resistant to spontaneous inactivation of their proteolytic activity than are dilute solutions, with the proteinase stability in the order of stem bromelain>fruit bromelain approximately ananain. The proteolytic activity of concentrated bromelain solutions remains relatively stable for at least 1 week at room temperature, with minimal inactivation by multiple freeze-thaw cycles or exposure to the digestive enzyme trypsin. The relative stability of concentrated versus dilute bromelain solutions to inactivation under physiologically relevant conditions suggests that delivery of bromelain as a concentrated bolus would be the preferred method to maximize its proteolytic activity in vivo.

Bromelains↗
PubMed2005

Adherent biofilms in bacterial vaginosis.

OBJECTIVE: Bacterial vaginosis is a common infectious disorder. Although known since ancient times, little progress has occurred in identifying causal factors. Our aims were to study the bacterial community structure and the spatial organization of microbiota on the epithelial surfaces of vaginal biopsy specimens. METHODS: We investigated the composition and spatial organization of bacteria associated with the vaginal epithelium in biopsy specimens from 20 patients with bacterial vaginosis and 40 normal premenopausal and postmenopausal controls using a broad range of fluorescent bacterial group-specific rRNA-targeted oligonucleotide probes. RESULTS: Bacterial vaginosis was associated with greater occurrence and higher concentrations of a variety of bacterial groups. However, only Gardnerella vaginalis developed a characteristic adherent biofilm that was specific for bacterial vaginosis. CONCLUSION: A biofilm comprised of confluent G vaginalis with other bacterial groups incorporated in the adherent layer is a prominent feature of bacterial vaginosis. LEVEL OF EVIDENCE: II-2.

Adolescent↗
PubMed2005

Piroxicam treatment of IL-10-deficient mice enhances colonic epithelial apoptosis and mucosal exposure to intestinal bacteria.

Treatment with the nonsteroidal anti-inflammatory drugs piroxicam or sulindac was recently shown to accelerate the development of colitis in interleukin (IL)-10-deficient (IL-10) mice. Although NSAIDs have been hypothesized to decrease the barrier function of the intestinal epithelium, the mechanism by which this accelerates colitis in IL-10 mice is not well understood. In this study, the effects of piroxicam on the colonic mucosa of IL-10 C57BL/6 mice were evaluated histologically. The effect of piroxicam on intestinal epithelial cells in vitro was assessed using colorimetric and fluorescent assays for cell viability and apoptotic cell death. Interactions of intestinal bacteria with the colonic mucosa were evaluated by rRNA-directed fluorescence in situ hybridization. In vivo treatment of C57BL/6 IL-10 mice with oral piroxicam markedly enhanced apoptosis of colonic epithelium and resulted in focal erosion of the mucosal surface, enhanced bacterial adhesion and invasion, and accelerated the development of colitis. In vitro, piroxicam induced apoptosis of CT26 murine intestinal epithelial cells in a dose-dependent fashion. Piroxicam-induced apoptosis of CT26 cells could not be prevented by addition of exogenous IL-10; however, IL-10 did significantly enhance their rate of proliferation. Thus, exposure to piroxicam enhances intestinal epithelial apoptosis both in vitro and in vivo and facilitates adhesion and invasion of intestinal bacteria into mucosal tissues in vivo. The role of IL-10 in this process requires further study. These studies support the hypothesis that increased exposure of mucosal cells to intestinal bacteria may lead to development of intestinal inflammation in IL-10 or other genetically susceptible individuals.

Animals↗
PubMed2005

Spatial organization and composition of the mucosal flora in patients with inflammatory bowel disease.

The composition and spatial organization of the mucosal flora in biopsy specimens from patients with inflammatory bowel disease (IBD; either Crohn's disease or ulcerative colitis), self-limiting colitis, irritable-bowel syndrome (IBS), and healthy controls were investigated by using a broad range of fluorescent bacterial group-specific rRNA-targeted oligonucleotide probes. Each group included 20 subjects. Ten patients who had IBD and who were being treated with antibiotics were also studied. Use of nonaqueous Carnoy fixative to preserve the mucus layer was crucial for detection of bacteria adherent to the mucosal surface (mucosal bacteria). No biofilm was detectable in formalin-fixed biopsy specimens. Mucosal bacteria were found at concentrations greater than 10(9)/ml in 90 to 95% of IBD patients, 95% of patients with self-limiting colitis, 65% of IBS patients, and 35% of healthy controls. The mean density of the mucosal biofilm was 2 powers higher in IBD patients than in patients with IBS or controls, and bacteria were mostly adherent. Bacteroides fragilis was responsible for >60% of the biofilm mass in patients with IBD but for only 30% of the biofilm mass in patients with self-limiting colitis and <15% of the biofilm mass in patients with IBS. In contrast, bacteria which positively hybridized with the probe specific for Eubacterium rectale-Clostridium coccoides accounted for >40% of the biofilm in IBS patients but for <15% of the biofilm in IBD patients. In patients treated with (5-ASA) or antibiotics, the biofilm could be detected with 4,6-diamidino-2-phenylindole but did not hybridize with fluorescence in situ hybridization probes. A Bacteroides fragilis biofilm is the main feature of IBD. This was not previously recognized due to a lack of appropriate tissue fixation. Both 5-ASA and antibiotics suppress but do not eliminate the adherent biofilm.

Adult↗
PubMed2004

Postnatal thymus transplantation with immunosuppression as treatment for DiGeorge syndrome.

Complete DiGeorge syndrome is a fatal congenital disorder characterized by athymia, hypoparathyroidism, and heart defects. Less than half of patients are 22q11 hemizygous. The goal of this study was to assess if immune suppression followed by postnatal thymus transplantation would lead to T-cell function in 6 infant patients who had host T cells at the time of transplantation. All infants had fewer than 50 recent thymic emigrants (CD3(+)CD45RA(+)CD62L(+)) per cubic millimeter (mm(3)) and all had some proliferative response to the mitogen phytohemagglutinin. Four infants had rash, lymphadenopathy, and oligoclonal populations of T cells in the periphery. Five of 6 patients are alive at the follow-up interval of 15 months to 30 months. The 5 surviving patients developed a mean of 983 host CD3(+) T cells/mm(3) (range, 536/mm(3)-1574/mm(3)), a mean of 437 recent thymic emigrants/mm(3) (range, 196/mm(3)-785/mm(3)), and normal proliferative responses to phytohemaglutinin (follow-up from day 376 to day 873). The TCR repertoire became polyclonal in patients who presented with oligoclonal T cells. All patients had thymopoiesis on allograft biopsy. Postnatal thymus transplantation after treatment with Thymoglobulin shows promise as therapy for infants with complete DiGeorge syndrome who have significant proliferative responses to mitogens or who develop rash, lymphadenopathy, and oligoclonal T cells.

DiGeorge Syndrome↗
PubMed2004

Corticosteroids regulate epithelial cell differentiation and Hassall body formation in the human thymus.

The presence of characteristic epithelial swirls called Hassall bodies within the human thymic medulla has been used as an indicator of ongoing or recent thymopoiesis. We present a case where Hassall bodies were present in the absence of current or past thymopoiesis. The patient had been treated with corticosteroids for presumed asthma before his diagnosis of X-linked SCID. Two other cases of nonimmunodeficient patients treated with high-dose corticosteroids had markedly increased numbers of thymic Hassall bodies. To determine whether corticosteroid treatment induces thymic epithelial (TE) differentiation to form Hassall bodies, mAbs reactive with specific cytokeratins (CKs), filaggrin, and involucrin were used to define distinct stages of TE cell differentiation. Treatment of primary TE monolayers with hydrocortisone in vitro induced expression of involucrin and high-molecular-mass CKs that are characteristic of TE differentiation. Treatment of thymic organ cultures with hydrocortisone induced both medullary and subcapsular cortical TE cells to express CK6, a differentiation marker that is normally expressed only by Hassall bodies in vivo. These experimental studies combined with the case observations indicate that exogenous corticosteroids can regulate terminal differentiation of TE cells both in vitro and in vivo. Thus, the presence of Hassall bodies in thymus from corticosteroid-treated patients cannot be taken as an absolute indication of previous thymopoiesis. Because corticosteroids are also made within the thymus under normal physiologic conditions, these studies support the hypothesis that endogenous corticosteroids may play a role in normal TE differentiation and Hassall body formation in vivo.

Adrenal Cortex Hormones↗
PubMed2004

Histologic and molecular assessment of human thymus.

Most work describing the histopathology of normal human thymus has focused on pediatric thymus because of tissue availability and high thymopoietic activity. However, pathologic examination of the thymus can provide information about immune status that is relevant to the clinical care of patients of all ages. Understanding age-related changes in the relative abundance and composition of anatomic compartments within the thymus is critical for evaluation of the thymus in normal adults and patients with diseases that affect the thymus. The purpose of this review is to acquaint diagnostic pathologists with some of the newer histologic, flow cytometric, and molecular techniques for assessment of non-neoplastic thymus. Diagnostic criteria are presented for assessment of thymic function and for determining the mechanisms underlying thymic hyperplasia. Accurate assessment of thymic function is critical for the diagnosis and treatment of patients with complete DiGeorge syndrome and can complement the clinical care of patients with a variety of disorders that affect the immune system.

Aged↗
PubMed2004

Abnormal development of thymic dendritic and epithelial cells in human X-linked severe combined immunodeficiency.

The X-linked form of severe combined immunodeficiency (X-SCID) is caused by mutations in the common cytokine receptor gamma chain and results in lack of T and NK cells and defective B cells. Without immune reconstitution, X-SCID patients typically die from infection during infancy. This report describes thymic epithelial (TE), lymphocyte, and dendritic cell (DC) differentiation in the thymic microenvironment of seven X-SCID patients who died before or after treatment for their immunodeficiency. X-SCID thymus consisted predominately of TE cells without grossly evident corticomedullary distinction. CD3+ and CD1a+ developing T cells and CD83+ thymic DC were reduced >50-fold when compared to age- and gender-matched control thymus (P < 0.001). TE expression of epithelial differentiation markers CK14, involucrin, and high molecular weight cytokeratins also differed in X-SCID versus normal thymus. These histopathologic findings indicate that in addition to T cells, thymic DC development and differentiation of TE cells are also abnormal in X-SCID.

Bone Marrow Transplantation↗
PubMed2004

Pathology of parainfluenza virus infection in patients with congenital immunodeficiency syndromes.

Infection with parainfluenza virus typically produces a mild, self-limited upper respiratory infection. However, parainfluenza infections have become increasingly recognized as a source of severe morbidity and mortality in immunocompromised patients. In this retrospective study we identified 6 patients with congenital immunodeficiency and positive respiratory cultures for parainfluenza virus who died and underwent complete autopsy. Tissues obtained at autopsy were studied using hematoxylin and eosin-stained sections, immunoperoxidase staining for parainfluenza virus, and in selected cases, electron microscopy. All 6 patients exhibited typical cytopathic effects of parainfluenza virus, including giant cell formation, in lung and/or bronchial tissues. Parainfluenza virus infection was also documented by giant cell formation and immunohistochemistry in the pancreas (in 3 of 6 patients) and the kidney or bladder (in 2 of 4 patients). Anti-parainfluenza antibody also specifically reacted with cells in the gastrointestinal tract (in 2 of 4), spleen (in 4 of 6), thymus and/or lymph nodes (in 4 of 4), and small blood vessels in various organs (in 4 of 6). Pancreatic, bladder, colon, and thymic epithelial cell lines were susceptible to experimental infections with clinical isolates of parainfluenza virus type 3 in vitro. Parainfluenza virus infection was serious in patients with congenital immunodeficiencies, contributing directly to death in 5 of the 6 patients studied. Because this virus is capable of infecting tissues in the gastrointestinal and urinary systems as well as in the respiratory tract, body secretions and fluids from each of these locations should be considered potentially infectious.

Cell Line↗
PubMed2004

Proteolytic activity and immunogenicity of oral bromelain within the gastrointestinal tract of mice.

Bromelain is a mixture of proteinases derived from pineapple stem that is marketed by health food stores as a "digestive aid". A number of studies suggest that bromelain may also have anti-inflammatory activity in vivo, including an anecdotal report describing potential efficacy in inflammatory bowel disease. We and others have previously shown that proteolytically active bromelain removes certain cell surface molecules and affects leukocyte migration, activation, and production of cytokines and inflammatory mediators in vitro. The purpose of this study was to determine whether ingested bromelain retains proteolytic activity within the murine gastrointestinal tract in vivo. The proteolytic activity of bromelain was determined in vitro using model substrates or immunofluorescence assays after administration of various doses and formulations orally to mice. Immune responses against bromelain were detected by enzyme immunoassays. When formulated in antacid, oral bromelain retained substantial proteolytic activity throughout the gastrointestinal tract. Bromelain concentrations within the colon were dependent on both dose and formulation and were sufficient to remove bromelain-sensitive molecules from both leukocytes and colon epithelial cells. Peak activity in the stool was observed 4 h after oral dosing. Although anti-bromelain IgG was detected in both serum and stool after long-term oral therapy, these antibodies did not prevent bromelain proteolytic activity within the gastrointestinal tract. These studies demonstrate that bromelain enzymes can remain intact and proteolytically active within the murine gastrointestinal tract. They provide further support for the hypothesis that oral bromelain may potentially modify inflammation within the gastrointestinal tract via local proteolytic activity within the colonic microenvironment.

Administration, Oral↗
PubMed2004

Complete DiGeorge syndrome: development of rash, lymphadenopathy, and oligoclonal T cells in 5 cases.

BACKGROUND: Five patients with DiGeorge syndrome presented with infections, skin rashes, and lymphadenopathy after the newborn period. T-cell counts and function varied greatly in each patient. Initial laboratory testing did not suggest athymia in these patients. OBJECTIVE: The purpose of this study was to determine whether the patients had significant immunodeficiency. METHODS: Research testing of peripheral blood included immunoscope evaluation of T-cell receptor beta variable gene segment repertoire diversity, quantification of T-cell receptor rearrangement excision circles, and detection of naive T cells (expressing CD45RA and CD62L). RESULTS: The patients were classified as having DiGeorge syndrome on the basis of syndromic associations and heart, parathyroid, and immune abnormalities. Immunoscope evaluation revealed that the T-cell repertoires were strikingly oligoclonal in all patients. There were few recent thymic emigrants, as indicated by the very low numbers of naive T cells (<50/mm(3)) and the absence of T-cell receptor rearrangement excision circles. These studies showed that all 5 patients were athymic. Two patients died, one from infection. No thymus was found during the complete autopsy performed on one patient. CONCLUSION: Patients with DiGeorge syndrome, skin rash, and lymphadenopathy should undergo analysis of naive T-cell numbers and of T-cell receptor beta variability segment repertoire to determine whether they are athymic, even if they have T cells with mitogen responsiveness. It is important for physicians to realize that patients with complete DiGeorge syndrome remain profoundly immunodeficient after development of these atypical features (rash, lymphadenopathy, and oligoclonal T cells). Prompt diagnosis is necessary for appropriate management.

Child, Preschool↗
PubMed2004

Thymic transplantation for complete DiGeorge syndrome: medical and surgical considerations.

BACKGROUND/PURPOSE: Complete DiGeorge syndrome results in the absence of functional T cells. Our program supports the transplantation of allogeneic thymic tissue in infants with DiGeorge syndrome to reconstitute immune function. This study reviews the multidisciplinary care of these complex infants. METHODS: From 1991 to 2001, the authors evaluated 16 infants with complete DiGeorge syndrome. All infants received multidisciplinary medical and surgical support. Clinical records for the group were reviewed. RESULTS: Four infants died without receiving a thymic transplantation, and 12 children survived to transplantation. The mean age at time of transplantation was 2.7 months (range, 1.1 to 4.4 months). All 16 infants had significant comorbidity including congenital heart disease (16 of 16), hypocalcemia (14 of 16), gastroesophageal reflux disease or aspiration (13 of 16), CHARGE complex (4 of 16), and other organ involvement (14 of 16). Nontransplant surgical procedures included central line placement (15 of 16), fundoplication or gastrostomy (10 of 16), cardiac repair (10 of 16), bronchoscopy or tracheostomy (6 of 16), and other procedures (12 of 16). Complications were substantial, and 5 of the 12 transplanted infants died of nontransplant-related conditions. All surviving infants have immune reconstitution, with follow-up from 2 to 10 years. CONCLUSIONS: Although the transplantation of thymic tissue can restore immune function in infants with complete DiGeorge syndrome, these children have substantial comorbidity. Care of these children requires coordinated multidisciplinary support.

DiGeorge Syndrome↗
PubMed2003

Thymus transplantation in complete DiGeorge syndrome: immunologic and safety evaluations in 12 patients.

Complete DiGeorge syndrome is a fatal condition in which infants have no detectable thymus function. The optimal treatment for the immune deficiency of complete DiGeorge syndrome has not been determined. Safety and efficacy of thymus transplantation were evaluated in 12 infants with complete DiGeorge syndrome who had less than 20-fold proliferative responses to phytohemagglutinin. All but one had fewer than 50 T cells/mm3. Allogeneic postnatal cultured thymus tissue was transplanted. T-cell development was followed by flow cytometry, lymphocyte proliferation assays, and T-cell receptor Vbeta (TCRBV) repertoire evaluation. Of the 12 patients, 7 are at home 15 months to 8.5 years after transplantation. All 7 survivors developed T-cell proliferative responses to mitogens of more than 100 000 counts per minute (cpm). By one year after transplantation, 6 of 7 patients developed antigen-specific proliferative responses. The TCRBV repertoire showed initial oligoclonality that progressed to polyclonality within a year. B-cell function developed in all 3 patients tested after 2 years. Deaths were associated with underlying congenital problems. Risk factors for death included tracheostomy, long-term mechanical ventilation, and cytomegalovirus infection. Adverse events in the first 3 months after transplantation included eosinophilia, rash, lymphadenopathy, development of CD4-CD8- peripheral T cells, elevated serum immunoglobulin E (IgE), and possible pulmonary inflammation. Adverse events related to the immune system occurring more than 3 months after transplantation included thrombocytopenia in one patient and hypothyroidism and alopecia in one other patient. Thymic transplantation is efficacious, well tolerated, and should be considered as treatment for infants with complete DiGeorge syndrome.

Child↗