Postthymectomy wasting associated with autoimmune phenomena. I. Antiglobulin-positive anemia in A and C57BL-6 Ks mice.

1. Mice of A and C(57)BL/6 Ks strains, thymectomized at birth acquire wasting disease in 84.1% (A) and 77.1% (C(57)BL/6 Ks) of the cases. There is no sex predelection. 2. Anemia in these animals is characterized by shortened red cell survival and increased fragility to hypotonic salt solutions. Among thymectomized A mice reticulocytosis is absent and extramedullary hematopoiesis is found in the spleen in the presence of bone marrow hypoplasia for the erythroid and lymphocyte series. 3. Positive antiglobulin tests of the red cells were observed in all the thymectomized C(57)BL/6 Ks (7/7) and 71.2% of the A strains (62/87). Normal mice do not show positive Coombs' tests. 4. The globulin coat on the A strain consists of IgM, whereas beta(1C) and IgG are not detectable. By contrast, red cell coats of NZB mice developing spontaneous autoimmune hemolytic anemia show IgM and beta(1C), but these erythrocytes do not react with anti-gamma chain antibodies. Another difference in the globulin coats of the two types of erythrocytes is that the IgM on NZB red cells has available light chain determinants but these are apparently hidden or absent in the case of sensitized erythrocytes. The difference in antibody coating, association with a component of complement in one but not the other, suggests a different mechanism for the immune surface phenomenon in each instance. 5. Anemia in NZB mice is associated with reticulocytosis while that in thymectomized A mice is not. 6. Thymectomy appears to initiate a chain of events leading to a series of autoimmune phenomena which may be due to alteration in host response consequent to loss of thymic tissue and thymic dependent functions or alternatively to infection to which increased susceptibility exists as a result of thymic extirpation.

Evidence that singlet oxygen-induced human T helper cell apoptosis is the basic mechanism of ultraviolet-A radiation phototherapy.

Ultraviolet A (UVA) irradiation is effectively used to treat patients with atopic dermatitis and other T cell mediated, inflammatory skin diseases. In the present study, successful phototherapy of atopic dermatitis was found to result from UVA radiation-induced apoptosis in skin-infiltrating T helper cells, leading to T cell depletion from eczematous skin. In vitro, UVA radiation-induced human T helper cell apoptosis was mediated through the FAS/FAS-ligand system, which was activated in irradiated T cells as a consequence of singlet oxygen generation. These studies demonstrate that singlet oxygen is a potent trigger for the induction of human T cell apoptosis. They also identify singlet oxygen generation as a fundamental mechanism of action operative in phototherapy.

The 55-kD tumor necrosis factor receptor on human keratinocytes is regulated by tumor necrosis factor-alpha and by ultraviolet B radiation.

In previous studies we showed that cultured human keratinocytes expressed the 55-kD TNF receptor (TNFR) and that its expression the important for TNF alpha-mediated upregulation of intercellular adhesion molecule-1 (ICAM-1) expression on keratinocytes. Because factors that either reduce or enhance TNFR expression are likely to have a major impact on the biological effects of TNF alpha on keratinocytes, these studies were conducted to determine the factors that regulate its expression on keratinocytes. Using reverse transcriptase polymerase chain reaction, human keratinocytes were shown to lack 75-kD TNFR expression, indicating that TNF responsiveness of human keratinocytes critically depended on regulation of 55-kD TNFR expression. Human keratinocyte 55-kD TNFR surface and mRNA expression was found to be regulated in vitro by recombinant human (rh) TNF alpha. Stimulation of keratinocytes with rhTNF alpha initially decreased, but later increased, 55-kD TNFR surface expression. This biphasic modulation of 55-kD TNFR surface expression was associated with concomitant changes in 55-kD TNFR mRNA expression. Ultraviolet B (UVB) radiation, a well-known inducer of synthesis and secretion of TNF alpha by human keratinocytes, was found to mimic TNF alpha-induced modulation of 55-kD TNFR surface and mRNA expression via a TNF alpha-mediated autocrine regulatory mechanism. Production of soluble 55-kD TNFR by human keratinocytes remained unaffected by TNF alpha stimulation or UVB irradiation. These studies provide clear evidence that membrane expression of the human 55-kD TNFR may be regulated in human keratinocytes by the ligand itself: TNF alpha. Since in previous studies UVB irradiation transiently inhibited TNF alpha-induced human keratinocyte ICAM-1 expression, it is proposed that UVB radiation-induced biphasic modulation of human keratinocyte 55-kD TNFR expression may affect the capacity of these cells to respond to TNF alpha.

Regulation of cell growth and cyclin D1 expression by the constitutively active FRAP-p70s6K pathway in human pancreatic cancer cells.

The FRAP-p70s6K signaling pathway was found to be constitutively phosphorylated/active in MiaPaCa-2 and Panc-1 human pancreatic cancer cells and a pancreatic cancer tissue sample as judged by the retarded electrophoretic mobility of the two major FRAP downstream targets, p70s6K and 4E-BP1. Treatment of cells with rapamycin, a selective FRAP Inhibitor, inhibited basal p70s6K kinase activity and induced dephosphorylation of p70s6K and 4E-BP1. Moreover, rapamycin inhibited DNA synthesis as well as anchorage-dependent and -independent proliferation in MiaPaCa-2 and Panc-1 cells. Finally, rapamycin strikingly inhibited cyclin D1 expression in pancreatic cancer cells. Thus, inhibitors of the constitutively active FRAP-p70s6K pathway may provide a novel therapeutic approach for pancreatic cancer.

Analysis of the production of soluble ICAM-1 molecules by human cells.

The adhesion molecule intercellular adhesion molecule-1 (ICAM-1), in addition to its membrane-associated form (mICAM-1), also exists as a soluble form (sICAM-1). sICAM-1 is capable of binding to lymphocyte function associated antigen-1 (LFA-1) molecules, and production of sICAM-1 is therefore thought to have immunomodulatory consequences. The present study, which employed normal human keratinocytes as a model for sICAM-1-producing cells, was conducted to determine the mechanism responsible for the production of sICAM-1 and to develop a strategy for specific inhibition of sICAM-1 production. Stimulation of keratinocytes with recombinant human gamma-interferon (rhIFN-gamma) induced both expression of mICAM-1 and production of sICAM-1. Western blot analysis revealed that keratinocyte-derived sICAM-1, compared to mICAM-1, had a smaller molecular size, approximately a 7-kD difference. Neither by Northern blot analysis nor by reverse-transcriptase polymerase chain reaction (RT-PCR) was any evidence for alternatively spliced ICAM-1 mRNA obtained. Addition of the protease inhibitors iodoacetamide and E-64, however, inhibited the production of sICAM-1 in a dose-dependent manner. The involvement of proteolytic cleavage in the production of sICAM-1 was corroborated in minimal peptide protection assays, in which minimal peptides covering the potential cleavage site of ICAM-1 were added to sICAM-1-producing keratinocytes. One of these peptides, ICAM cleavage inhibitory peptide (ICAM-CIP), inhibited the production of sICAM-1 without affecting mICAM-1 expression. These studies demonstrate that sICAM-1 production in human keratinocytes is due to proteolytic cleavage, and that the oligopeptide ICAM-CIP may specifically inhibit this mechanism. The capacity of ICAM-CIP to selectively prevent production of sICAM-1 may be useful for the development of novel therapeutic approaches relevant for the management of inflammation and cancer.

Involvement of cytokines, DNA damage, and reactive oxygen intermediates in ultraviolet radiation-induced modulation of intercellular adhesion molecule-1 expression.

By virtue of its capacity to serve as a counter-receptor for lymphocyte function-associated antigen-1, intercellular adhesion molecule-1 (ICAM-1) plays a pivotal role in generation and maintenance of immunologic/inflammatory skin diseases by mediating leukocyte/keratinocyte adhesion. Ultraviolet radiation (UVR) may exert both antiinflammatory effects (e.g., UV phototherapy) and proinflammatory effects (e.g., triggering of photosensitive skin diseases) on human skin. Recent evidence indicates that UVR-induced changes of keratinocyte ICAM-1 expression constitute the molecular basis for these ambivalent properties of UVR, as UVR is able to exert two separate and even opposite effects on ICAM-1 expression. As an antiinflammatory effect, UVR may inhibit cytokine-induced up-regulation of keratinocyte ICAM-1 expression, whereas induction of ICAM-1 expression by UVR represents a proinflammatory activity. This latter effect is mediated by an autocrine mechanism involving interleukin (IL)-1 alpha. In this autocrine system, UVR exposure of human keratinocytes leads to the release of IL-1 alpha, which in turn up-regulates the expression of IL-1 receptor type 1 molecules on the keratinocyte surface, thereby increasing the sensitivity of these cells toward IL-1 alpha. As a consequence, irradiated keratinocytes are capable of responding to endogenously produced IL-1 alpha by increasing ICAM-1 expression. Modulation of keratinocyte ICAM-1 expression after UVR exposure may be observed after both short-wave UVR (UVB; 280-320 nm) and long-wave UVR (UVA1; 340-400 nm). The photobiologic mechanisms underlying UVB versus UVA1 radiation-induced ICAM-1 modulation have been found to differ. Although not completely delineated, UVB radiation-induced modulation of ICAM-1 expression appears to be mediated via the induction of DNA damage, whereas UVA1 radiation effects involve the generation of reactive oxygen intermediates.

Interleukin-10 production by cultured human keratinocytes: regulation by ultraviolet B and ultraviolet A1 radiation.

Keratinocytes are the primary cellular target for ultraviolet radiation in human skin, and ultraviolet radiation-induced therapeutical effects may thus be mediated by keratinocyte-derived, antiinflammatory mediators. Interleukin-10 is capable of exerting antiinflammatory effects by virtue of its capacity to suppress the production of interferon-gamma. The present study therefore assessed the ability of cultured human keratinocytes to produce interleukin-10 following ultraviolet irradiation. Exposure of long-term cultured normal human keratinocytes to ultraviolet B (280-320 nm) or to ultraviolet A1 (340-400 nm) radiation caused a time- and dose-dependent induction of interleukin-10 mRNA expression and interleukin-10 protein secretion, with ultraviolet A1 radiation being the strongest stimulus. Ultraviolet radiation-induced interleukin-10 production by normal human keratinocytes was enhanced by a factor of two, when cells were cultured in high- rather than low-calcium medium. Neither addition of the ultraviolet radiation-inducible cytokines tumor necrosis factor-alpha or interleukin-1 alpha to unirradiated keratinocytes nor presence of their respective neutralizing antibodies in cultures of irradiated keratinocytes induced or inhibited interleukin-10 synthesis. Modulation of eicosanoid production by addition of prostaglandin E2 to keratinocyte cultures or disturbance of cyclooxygenase activity by indomethacin did not affect interleukin-10 production in resting or irradiated cells. These studies demonstrate that cultured human keratinocytes are capable of producing interleukin-10. Human keratinocyte interleukin-10 production is dependent on the differentiation state of the cell and induced by ultraviolet B and, in particular, ultraviolet A1 radiation exposure. This novel property of ultraviolet radiation may account at least in part for the efficacy of phototherapy in inflammatory skin diseases.

Neurotrophin-4 production by human epidermal keratinocytes: increased expression in atopic dermatitis.

Chronic inflammatory conditions of human skin, such as prurigo lesions of atopic dermatitis, are characterized clinically by intense pruritus and histologically by increased innervation. Regulation of skin innervation is thought to depend on neurotrophic factors. In this study, human skin cells were identified as a source of neurotrophins. Cultured keratinocytes expressed neurotrophin-4, whereas dermal fibroblasts expressed neurotrophin-3. In vitro stimulation with interferon-gamma, a marker cytokine for atopic eczema, induced keratinocyte neurotrophin-4 production, which was able to support growth of a neuroglioblastoma-derived cell line. In vivo, immunohistochemistry of human skin for neurotrophins showed neurotrophin-4 staining in the epidermal layer and neurotrophin-3 staining in the dermal compartment. Neurotrophin-4 but not neurotrophin-3 expression was markedly increased in interferon-gamma-injected skin. Prurigo lesions of atopic dermatitis skin were characterized by intense epidermal staining for neurotrophin-4, suggesting a pathophysiologic role for this neurotrophin in the increased innervation characteristic for these skin lesions. This study demonstrates differential expression and regulation of neurotrophins in human skin. It also identifies keratinocyte-derived neurotrophin-4 as a possible link between the immune and the nerve system of human skin.

Post-transcriptional regulation of interleukin-6 gene expression in human keratinocytes by ultraviolet B radiation.

Exposure to increasing doses (290-315 nm) of ultraviolet (UV) B radiation is thought to profoundly affect human health. Studies on the biologic and molecular effects of UVB radiation on human skin are therefore of particular interest. There is experimental and clinical evidence to assume that UVB radiation-induced local and systemic inflammatory reactions might be mediated at least in part by UVB-induced keratinocyte-derived interleukin (IL)-6. Previously, a UVB-induced increase of steady-state levels of IL-6 mRNA was found to be a prerequisite for keratinocyte IL-6 production after UVB irradiation. The present study was aimed at addressing the question of whether in vitro UVB irradiation would increase IL-6 mRNA expression in long-term cultured, normal human keratinocytes via transcriptional or post-transcriptional mechanisms. UVB exposure (0-100 J/m2) of keratinocytes increased low baseline expression levels of IL-6 mRNA in a time- and dose-dependent manner. Using nuclear run-on assays, transcription rates of the IL-6 gene in nuclei isolated from UVB-irradiated cells were found to be essentially identical to those seen in unirradiated cells, indicating that UVB light did not lead to increased transcription of the IL-6 gene. To determine a possible post-transcriptional mechanism in UVB-induced IL-6 mRNA expression, the effects of UVB irradiation on IL-6 mRNA stability were examined. To this end irradiated and unirradiated keratinocytes were treated with actinomycin D and subjected to Northern blot analysis to calculate IL-6 mRNA half-life. As compared with unirradiated cells, IL-6 mRNA stability was increased significantly (three- to four-fold) in UVB-irradiated cells, suggesting that UVB radiation upregulates IL-6 mRNA levels in human keratinocytes by increasing the stability of IL-6 transcripts. This is the first report indicating that UVB radiation at a physiologically relevant dose may affect gene expression in human cells at a post-transcriptional level.

Analysis of the mechanism of ultraviolet (UV) B radiation-induced prostaglandin E2 synthesis by human epidermoid carcinoma cells.

Stimulation of cultured human keratinocytes with interleukin (IL)-1 alpha is known to elicit prostaglandin (PG) E2 release. Ultraviolet (UV) B radiation induces keratinocyte PGE2 and cytokine production. The present study deals with the autocrine roles of UVB-induced, keratinocyte-derived cytokines IL-1 and tumor-necrosis-factor (TNF) alpha and their corresponding receptor molecules for UVB-induced PGE2 release. In vitro exposure of transformed human keratinocytes (KB cells) induced PGE2 production five- to eightfold. This increase was inhibited by 70%, if irradiated cells were cultured in presence of monoclonal antibody (MoAb) M4, which blocks IL-1 effects by binding to the type 1 IL-1 receptor (IL-1R). In contrast, MoAb M22, which blocks the type 2 IL-1R, had no significant effects. Addition of recombinant human TNF alpha to unirradiated KB cells resulted in five- to eightfold increased PGE2 synthesis, and this increase could be mimicked by stimulation of KB cells with MoAb htr-9, which exerts TNF alpha-like bioactivity by binding to the 55-kD TNF receptor (TNFR). UVB-induced PGE2 synthesis was blocked by 50% in the presence of neutralizing anti-TNF alpha-Ab, and was completely inhibited by addition of both anti-TNF alpha-Ab and MoAb M4. To elucidate a possible regulatory intracellular step in PGE2 synthesis, specific cyclooxygenase activity in KB cells was determined. Following UVB treatment, cyclooxygenase activity increased twofold, but remained unaltered, if irradiated KB cells were cultured in the presence of anti-TNF alpha-Ab plus MoAb M4. These studies indicate that keratinocyte-derived TNF alpha and IL-1 together mediate UVB-induced PGE2 release via specific cell surface receptors, and that one intracellular mechanism is an increased prostanoid-synthesizing capacity of irradiated cells.

Analysis of the cytokine pattern expressed in situ in inhalant allergen patch test reactions of atopic dermatitis patients.

In the present study, we examined the cytokine pattern expressed in situ during the development of eczematous reactions that had been provoked in atopic dermatitis patients by patch testing with house dust mite allergen. In 24-h house dust mite allergen patch test reactions, expression of interleukin (IL)-4 mRNA and IL-2 mRNA increased, but interferon (IFN)-gamma mRNA did not, as compared with control skin. In 48-h inhalant allergen patch test reactions, however, IFN-gamma mRNA and IL-2 mRNA expression were increased above levels observed in control skin, whereas IL-4 mRNA expression was decreased below background levels. These data demonstrate that a switch from a Th2-like to a Th1-like cytokine response occurs in inhalant allergen patch tests of atopic dermatitis patients. This biphasic pattern was specific to inhalant allergen patch test reactions, as it was not observed in irritant reactions in the same patient. IFN-gamma production by T cells may be induced by the cytokine IL-12. In the present study, up-regulation of IFN-gamma mRNA expression in inhalant allergen patch test reactions was preceded by an increased expression of the p35 subunit of IL-12 mRNA. These observations suggest that increased IL-12 expression may contribute to the observed switch of the in situ cytokine secretion pattern. Further studies are necessary to determine whether a similar biphasic pattern of cytokine expression is also operative in the initiation phase of atopic eczema.

Interleukin-1 receptors type I and type II are differentially regulated in human keratinocytes by ultraviolet B radiation.

Since regulation of keratinocyte IL-1 receptor expression is likely to have a major impact on the biologic effects of IL-1 on epidermal cells, we examined expression, regulation, and function of IL-1R in cultured human keratinocytes. By reverse transcriptase polymerase chain reaction, human keratinocytes were shown to express IL-1 receptor type I (IL-1RI) and IL-1 receptor type II (IL-1RII). Human keratinocyte IL-1RI mRNA expression was dependent on the differentiation state of the cell and was regulated by ultraviolet B (UVB) radiation, which initially decreased but later increased IL-1RI expression. This UVB-induced biphasic modulation of IL-1RI expression was mediated by an autocrine mechanism involving endogenously produced IL-1alpha and IL-1RI. Increased expression of IL-1RI in UVB-irradiated or IL-1alpha-stimulated keratinocytes was functionally important, because it endowed these cells with the capacity to upregulate expression of the intercellular adhesion molecule (ICAM)-1 upon IL-1alpha stimulation. Keratinocyte IL-1RII expression was regulated by UVB irradiation in an inverse manner. Significant and rapid upregulation of IL-1RII was observed within 1 h after UVB irradiation and gradually decreased to background levels within 24 h. Inverse regulation of IL-1RII versus IL-1RI was associated with opposite functions, because blocking of IL-1RII enhanced IL-1alpha effects on induction of ICAM-1 expression. These studies demonstrate that IL-1 responsiveness of UVB-irradiated keratinocytes critically depends on regulation of IL-1RI expression and that IL-1RII serves as a "decoy" receptor for IL-1, limiting rather than promoting IL-1-mediated effects.

Allergen specificity of skin-infiltrating T cells is not restricted to a type-2 cytokine pattern in chronic skin lesions of atopic dermatitis.

The majority of allergen-specific T cells derived from inhalant allergen patch test lesions in patients with atopic dermatitis were previously found to produce a restricted type-2 cytokine pattern. Recent studies, however, have revealed that in chronic eczematous skin lesions of patients with atopic dermatitis, expression of the type-1 cytokine interferon-gamma predominates. To evaluate cytokine production by allergen-specific T cells in chronic atopic dermatitis, we established house dust mite (Dermatophagoides pteronyssinus)-specific T-cell clones from the dermis of chronic skin lesions of sensitized adult patients with atopic dermatitis. Frequencies of skin-derived T cells proliferating in the presence of Dermatophagoides pteronyssinus were between one in 138 and one in 4255, indicating that only a minority of skin-infiltrating T cells are allergen specific. When these cells were analyzed for their capacity to produce interferon-gamma, the majority (71%) of these cells were found to express interferon-gamma mRNA and to secrete interferon-gamma protein, either alone or in combination with interleukin-4. Phenotypic analysis revealed that 15% of skin-infiltrating allergen-specific T cells were CD8+. No selection of Vbeta elements was detected in Dermatophagoides pteronyssinus-specific T-cell clones. These studies demonstrate that allergen specificity of skin-infiltrating T cells is not restricted to a type-2 cytokine pattern in lesional atopic dermatitis. The notion that the majority of allergen-specific, skin-infiltrating T cells are capable of producing interferon-gamma further supports the concept that interferon-gamma expression has major pathogenetic relevance for the chronic phase of atopic dermatitis.

High-dose ultraviolet A1 (UVA1), but not UVA/UVB therapy, decreases IgE-binding cells in lesional skin of patients with atopic eczema.

In order to further elucidate the mechanisms by which high-dose ultraviolet A1 (UVA1) therapy leads to improvement in patients with atopic eczema, we assessed skin sections from patients before and after high-dose UVA1 therapy (n = 5) or conventional UVA/UVB therapy (n = 4) for changes in Langerhans cells and mast cells expressing the high-affinity IgE receptor Fc epsilon RI and in surface-bound IgE by histochemical and immunohistochemical techniques. The two treatment groups exhibited different patterns of changes in the number of Fc epsilon RI+, CD1a+, and mast cells within the dermis: The density of both Langerhans cells and mast cells was decreased after high-dose UVA1 therapy, but not after UVA/UVB therapy. High-dose UVA1 and UVA/UVB therapy significantly increased the number of CD1a+ cells within the epidermis, but only high-dose UVA1 reduced the relative number of IgE+ intraepidermal Langerhans cells typically found in atopic eczema. Reduction of numbers of dermal Langerhans cells and mast cells, as well as relative numbers of intraepidermal IgE+ Langerhans cells, was closely linked to significant clinical improvement by high-dose UVA1, but not UVA/UVB therapy. These studies support the notion that IgE-binding cutaneous cells are involved in the pathogenesis of atopic eczema. We propose that UVA1 radiation exerts its effects in atopic eczema, at least in part, by inhibiting Langerhans cell migration out of the epidermis and, in particular, by reducing the number of IgE-bearing Langerhans cells and mast cells in the dermis.

Induction of interleukin-6 production by ultraviolet radiation in normal human epidermal keratinocytes and in a human keratinocyte cell line is mediated by DNA damage.

The sunburn reaction is the most common consequence of human exposure to ultraviolet radiation (UVR), and is mediated at least in part by interleukin-6 (IL-6). The aim of this study was to determine if DNA is a major chromophore involved in the induction of IL-6 following UV irradiation of a human epidermoid carcinoma cell line (KB), and of normal human epidermal keratinocytes. We first confirmed that IL-6 release was associated with enhanced levels of IL-6 mRNA transcripts. The wavelength dependence for IL-6 release was then investigated by irradiating the cells at defined wavelengths (254, 302, 313, 334, and 365 nm) with a monochromator. The maximum effect on IL-6 release was observed at 254 nm with only low levels of induction observed at wavelengths above 313 nm. The wavelength dependence for UV-induced IL-6 release was similar to that for DNA absorption or for the induction of cyclobutane pyrimidine dimers (CPD). To determine whether UV-induced DNA damage mediated IL-6 secretion, the role of CPD was investigated by treating keratinocytes with photosomes (photolyase encapsulated in liposomes) followed by photoreactivating light. This photoreversal procedure led to a reduction in the levels of the UVC-induced secretion of IL-6, which in normal human keratinocytes was unambiguously associated with repair of CPD. We conclude that the release of IL-6 from human keratinocytes following short-wave UVC and UVB irradiation is mediated by DNA damage and that CPD play an important role in this process.

Secretory phospholipase A2 in patients with infected pancreatic necroses in acute pancreatitis.

Secretory synovial-type PLA2 (sPLA2-II) in peripheral blood is known to be associated with systemic complications in patients with severe diseases. Being the pacemaking enzyme in eicosanoid synthesis, sPLA2-II is a mediator of the inflammatory response and plays a role in host defense against bacterial infection. We evaluated the clinical role of systemic sPLA2-II in bacterial infection of pancreatic necroses in severe acute pancreatitis. In 58 patients with acute pancreatitis, pancreatic and sPLA2-I and sPLA2-II were measured daily for the first 14 days of hospital treatment by a time-resolved fluoroimmunoassay. All 36 patients with necrotizing pancreatitis underwent regular fine needle aspiration (FNA) to monitor bacterial infection. In 10 patients, infected necroses were found on FNA and postoperative examination. On admission and at most days throughout the observation period, systemic sPLA2-II was significantly higher in patients with infected necroses than in patients with sterile necroses or interstitial pancreatitis. This difference was not found for sPLA2-I, but values were higher in necrotizing pancreatitis than in interstitial pancreatitis at the first 2 days of hospital treatment. If sPLA2-II was >300 ng/ml on 2 successive days within the first 4 days, infected necroses could be predicted with a sensitivity of 89%, a specificity of 88%, and a negative predictive value of 95%. Systemic sPLA2-II has the potential to identify patients at risk of bacterial infection of pancreatic necroses and its routine measurement may therefore, in combination with FNA, offer a valuable tool in monitoring patients with acute necrotizing pancreatitis.

Fungal infection in acute necrotizing pancreatitis.

BACKGROUND: Anecdotal reports suggest that patients with fungal infection of necrotizing pancreatitis (NP) have worse outcomes than those with bacterial infection. Our aim was to compare the clinical course and outcomes of patients with NP infected with fungal versus nonfungal organisms. STUDY DESIGN: Prospectively collected data on 57 patients with infected NP (1983-1995) were reviewed. RESULTS: Seven patients (12%) developed fungal infection, and 50 (88%) developed bacterial infection. Groups had similar mean ages (60 versus 63 years) and APACHE-II scores on admission (9 each). The cause of NP was ERCP-induced in 3 of 7 with fungal infection versus 3 of 50 with bacterial infection. Patients with fungal infection had been treated with a mean of 4 different antibiotics for a mean of 23 days, and 4 of 7 (57%) required mechanical ventilation preoperatively. In addition, postoperative ICU stays were longer (20 versus 10 days), as were total hospital stays (59 versus 41 days). Mortality was higher with fungal infection; 3 of 7 patients (43%) died versus 10 of 50 patients (20%). CONCLUSIONS: Although NP presents with similar initial severity, patients with fungal infection of NP tend to have a more complicated course and worse outcomes compared with those with bacterial infection. Low-dose antifungal prophylaxis should be added to early management of NP.

Induction of proinflammatory cytokines in human epidermoid carcinoma cells by in vitro ultraviolet A1 irradiation.

Ultraviolet radiation-induced expression of cytokines by keratinocytes is important for the pathogenesis of polymorphous light eruption (PLE). Because UVA1 radiation rather than UVB radiation might be a more important trigger for PLE, cells from the human epidermoid carcinoma cell line KB were exposed in vitro to UVA1 radiation (30 J/cm2) and subsequently analyzed for cytokine expression. Ultraviolet A1 irradiation induced tumor necrosis factor (TNF)-alpha and interleukin (IL)-8 expression in KB cells at the mRNA and protein level. Upregulation of cytokine mRNA levels followed a biphasic pattern. This effect was specific for TNF alpha and IL-8 because UVA1 radiation did not induce expression of IL-1 alpha or IL-6 in these cells. Ultraviolet A1 radiation-induced expression of intercellular adhesion molecule-1 in KB cells previously was found to depend on the thiol status of these cells. Therefore, KB cells were treated with DL-buthionine-[S,R]-sulfoximine (BSO), a specific inhibitor of de novo glutathione synthesis. Exposure of BSO-pretreated KB cells to UVA1 radiation significantly induced IL-1 alpha and IL-6 mRNA and protein expression. These studies demonstrate the capacity of UVA1 radiation to induce cytokine expression in human epidermoid carcinoma cells. This immunomodulatory effect may be mediated by thiol-status-dependent and -independent mechanisms.

Chronically ultraviolet-exposed human skin shows a higher mutation frequency of mitochondrial DNA as compared to unexposed skin and the hematopoietic system.

Normal ageing processes are associated with an accumulation of mutations within the mitochondrial (mt) DNA. The most frequent mutation is a 4977 base pair (bp) deletion known as common deletion. In order to test the hypothesis that chronically sun-exposed skin is characterized by an increased mutation frequency of mtDNA, the mutation frequency of the common deletion between skin and another replicating tissue (the hematopoietic system) and chronically sun-exposed versus sun-protected skin was compared in the same individuals. This was done by comparing the amount of mutated mtDNA molecules with the whole mitochondrial genome in the same specimen with a semiquantitative polymerase chain reaction method, thus allowing direct comparison of different tissues. In all skin specimens the common deletion could be observed. In contrast only 3 of 10 blood samples revealed detectable amounts of the common deletion. Comparison of sun-exposed versus sun-protected skin exhibited a higher content of the common deletion in sun-exposed skin in 7 of 10 individuals. Additionally, a hitherto undescribed mtDNA mutation was detected exclusively in human skin. These studies indicate that exposure of human skin to solar radiation leads to an accumulation of mtDNA mutations, possibly via oxidative damage, which may play an important role in photoageing.

Acoustic properties of particle/polymer composites for ultrasonic transducer backing applications.

The acoustic impedance and attenuation in composites made of particle fillers loaded in polymer matrices for transducer backing applications is investigated. The acoustic impedance of tungsten/vinyl composites was modeled, and an experimental matrix identifying variables that contribute to composite attenuation was established. The variable included the particle type, the particle size and volume fraction of a filler, the physical characteristics of the polymer matrix, and the processing route that determined the composite connectivity. Experimental results showed that with an increase in filler particle size or a decrease in volume fraction of filler, there is an increase in composite attenuation. Overall, the various types of filler, the polymer matrix, and the interface between the two contribute to attenuation in the composite, as confirmed by the acoustic properties and the microstructural analysis.