Long-term sleep deprivation in rats produces dramatic physiological changes including increase in energy expenditure, decrease in body weight, and death after 2-3 weeks. Despite several studies, the sleep deprivation syndrome remains largely unexplained. Here, to elucidate how prolonged sleep loss affects brain cells we used microarrays and screened the expression of>> 26 000 transcripts in the cerebral cortex. Rats were sleep deprived using the disk-over-water method for 1 week. Seventy-five transcripts showed increased expression in these animals relative to controls that had been spontaneously awake or sleep deprived for a few hours. Most of them were induced as a result of chronic sleep loss and not non-specific effects of the disk stimulation. They include transcripts coding for several immunoglobulins, stress response proteins (macrophage inhibitor factor-related protein 14, heat-shock protein 27, alpha-B-crystallin), minoxidil sulfotransferase, globins and cortistatin. Twenty-eight transcripts decreased their expression in long-term sleep-deprived rats. Sixteen of them were specifically decreased as a result of chronic sleep loss, including those coding for type I procollagen and dihydrolipoamide acetyltransferase. We also compared sleeping rats to short-term and long-term sleep-deprived rats, and found that acute and chronic sleep loss led to some differences at the molecular level. Several plasticity-related genes were strongly induced after acute sleep deprivation only, and several glial genes were down-regulated in both sleep deprivation conditions, but to a different extent. These findings suggest that sustained sleep loss may trigger a generalized inflammatory and stress response in the brain.

The central nervous system (CNS) presents both challenges and opportunities to researchers of redox biochemistry. The CNS is sensitive to oxidative damage during aging or disease; excellent transgenic models of specific neurodegenerative diseases have been created that reproduce oxidative stress components of the corresponding human disorder. Mouse models of familial amyotrophic lateral sclerosis (ALS) based on overexpressed mutant human Cu, Zn-superoxide dismutase (SOD1) are cases in point. These animals experience predictably staged, age-dependent motor neuron degeneration with profound cellular and biochemical damage to nerve fibers and spinal cord tissue. Severe protein and lipid oxidation occurs in these animals, apparently as an indirect consequence of protein aggregation or cytopathic protein-protein interactions, as opposed to aberrant redox catalysis by the mutant enzyme. Recent studies of G93A-SOD1 mice and rats suggest that oxidative damage is part of an unmitigated neuroinflammatory reaction, possibly arising in combination from mitochondrial dysfunction plus pathophysiologic activation of both astrocytes and microglia. Lesions to redox signal-transduction pathways in mutant SOD1+ glial cells may stimulate broad-spectrum upregulation of proinflammatory genes, including arachidonic acid-metabolizing enzymes [e.g., cyclooxygenase-II (COX-II) and 5-lipoxygenase (5LOX)]; nitric oxide synthase (NOS) isoforms; cytokines (particularly tumor necrosis factor alpha, TNF-alpha); chemokines; and immunoglobulin Fc receptors (FcgammaRs). The integration of these processes creates a paracrine milieu inconsistent with healthy neural function. This review summarizes what has been learned to date from studies of mutant SOD1 transgenic animals and demonstrates that the G93A-SOD1 mouse in particular is a robust laboratory for the study of neuroinflammation and redox biochemistry.

The B lymphocyte cell surface receptor CD22 is an adhesion molecule that can mediate binding to several leukocyte subsets. The first CD22 ligand to be identified was the receptor-linked phosphotyrosine phosphatase CD45, but several lines of evidence suggest that CD22 may interact with multiple counter receptors on adjacent lymphocytes. In the present work, we show that in addition to CD45, a soluble CD22-immunoglobulin fusion protein (CD22Rg) recognizes several other distinct lymphocyte sialoglycoproteins. CD22-mediated adhesion is dependent upon the presence of sialic acids on ligands. CD22Rg is observed to bind specifically to a 115-kDa sialoglycoprotein in COS cells transfected with an alpha-2,6-sialyltransferase cDNA, but not in COS cells transfected with unrelated cDNA clones, indicating that at least some CD22-mediated interactions require presentation of sialic acid in an alpha-2,6 linkage by CD22 ligands. In all cases, truncation of the side chain of sialic acids by mild periodate oxidation abolishes recognition by CD22Rg. Direct binding of CD22Rg to lymphoid cells also requires sialic acids and their side chains. Taken together, these observations indicate that CD22 is a sialic acid-binding lectin and may define a novel functional subset of immunoglobulin superfamily adhesion molecules.

CD22 beta is a glycoprotein found on the surface of B cells during restricted stages of development. It is believed to play a role in cell-cell interactions and B cell activation. The accompanying paper (Sgroi, D., Varki, A., Braesch-Andersen, S., and Stamenkovic, I. (1993) J. Biol. Chem. 268, 7011-7018) shows that CD22 beta recognizes multiple glycoproteins on the surfaces of T and B cells and that sialylation of these ligands is essential for binding. To identify the structure(s) of the sialylated oligosaccharide(s) recognized by CD22 beta, [3H]glucosamine-labeled glycoproteins were purified from Daudi cells by adsorption onto a CD22 beta recombinant immunoglobulin (CD22 beta Rg) chimera attached to protein A-Sepharose (PAS), and the N-linked oligosaccharides were released by peptide N-glycosidase F. These released oligosaccharides failed to bind to CD22 beta Rg-PAS under the conditions used initially to adsorb the glycoproteins, but their elution from a column of CD22 beta Rg-PAS was significantly retarded. Populations of oligosaccharides with different affinities could be identified by their order of elution. Specific sialidases were used to determine the content of alpha-2,3- and alpha-2,6-linked sialic acid in these different populations and their contribution to binding. Multiantennary oligosaccharides with one alpha-2,6-linked residue bound marginally, and those with two or more bound more tightly. alpha-2,3-Linked sialic acid residues were without effect. Binding did not require divalent cations and was abrogated by mild periodate oxidation of the outer side chain of sialic acid. No marked differences in size or fucose content were found between the populations of high and low affinity oligosaccharides. However, the low affinity population could be partially converted into higher affinity by treatment with beta-galactoside alpha-2,6 sialyltransferase and CMP-sialic acid. Thus, CD22 beta is a mammalian lectin that can recognize specific N-linked oligosaccharide structures containing alpha-2,6-linked sialic acids.

The anti-infectious effect of probiotics has recently been reported and one mechanism may be the non-specific stimulation of immunity. This study was performed to elucidate the influence of a probiotic formula on intestinal microflora and local immunity in healthy children. A follow-up formula containing viable bifidobacteria was given to seven healthy Japanese children (15 to 31 months old) for 21 days. During intake of the formula, the administered strain was detected in feces from five subjects (71%) and total fecal bifidobacteria slightly increased. Fecal levels of total IgA and anti-poliovirus IgA during intake of the formula were significantly higher than those before intake (P < 0.05). The increase in local IgA levels resulting from ingestion of the probiotic formula may contribute to enhancement of the mucosal resistance against gastrointestinal infections.

We studied the ability of the lipopolysaccharide (LPS) extracted from a vaccine strain of Francisella tularensis (LPS-Ft) to mimic LPSs from other gram-negative bacteria for activation of various murine cell types or to antagonize the effects of other LPSs. We found that activation of macrophages for the production of tumor necrosis factor alpha and NO, of pre-B lymphocytes for the expression of surface immunoglobulins, and of bone marrow cells for the expression of LPS-binding sites was either undetectable with LPS-Ft or required concentrations 100 to 1,000 times higher than for standard LPSs. Preexposure of macrophages to LPS-Ft also failed to trigger down-regulation of tumor necrosis factor alpha (desensitization) or up-regulation of NO responses to an endotoxin challenge. In contrast to other atypical LPSs, LPS-Ft was also unable to antagonize any of the endotoxin-induced cellular responses mentioned above, suggesting that this LPS does not interact with LPS receptors.

Studies performed at large metropolitan medical centers have reported an increasing incidence of idiopathic focal segmental glomerulosclerosis (FSGS) in adults. To determine whether a similar trend occurs in small urban and rural communities and to determine the role of race in these observations, we reviewed the patient records of all adults who underwent renal biopsies at our institution over the 20-year period from 1974 to 1994. The patients were grouped for analysis in 5-year intervals, 1975 to 1979, 1980 to 1984, 1985 to 1989, and 1990 to 1994, for the following diagnoses: FSGS, membranous nephropathy (MN), minimal change nephropathy (MCN), membranoproliferative glomerulonephritis (MPGN), immunoglobulin A (IgA) nephropathy, chronic glomerulonephritis, diabetic nephropathy, hypertensive nephrosclerosis, and chronic interstitial nephritis. Patients with secondary causes for these lesions were excluded. The relative frequency of FSGS increased from 13.7% during 1975 to 1979 to 25% during 1990 to 1994 (P < 0.05). The relative frequency of MN decreased from 38.3% during 1975 to 1979 to 14.5% during 1990 to 1994 (P < 0.01). There were no changes in the frequencies of MCN, MPGN, IgA nephropathy, chronic glomerulonephritis, diabetic nephropathy, hypertensive nephrosclerosis, or chronic interstitial nephritis over the 20-year period. However, there was a significant increase in the percentage of blacks with FSGS, from 0% in 1975 to 1979 to 22.6% in 1990 to 1994, and an increased percentage of Hispanics with FSGS, from 0% in 1975 to 1979 to 21.3% in 1990 to 1994 (P < 0.05). The modest increase in whites with FSGS did not reach statistical significance. The incidence of MN in blacks and whites decreased over the 20-year period. In the last 5 years, 15 patients per year had FSGS compared with 7 patients per year with MN (P < 0.05). No changes in age or sex between groups or over time accounted for these results. We conclude that FSGS is now diagnosed twice as often as MN and is the most common idiopathic glomerular disease at our hospital. Reasons for this increase include the emergence of FSGS in both Hispanics and blacks, with a modest increase of FSGS in whites. The increase in FSGS in the three most common races in our community suggests that factors other than genetic, perhaps environmental, have a role in the pathogenesis of FSGS.

The T-cell receptor is necessary and sufficient for recognition of peptides presented by major histocompatibility complex molecules. Other adhesion molecules, like CD4 or CD8, play an auxiliary role in antigen recognition by T cells. Here we analyse T-cell receptor (TCR) binding using a soluble rather than a cell-bound receptor molecule. A TCR-immunoglobulin chimaera is constructed with the variable and the first constant regions of both the TCR alpha- and beta-chains linked to the immunoglobulin light-chain constant regions. This soluble TCR is expressed, assembled and secreted as an alpha beta heterodimer by a myeloma cell line transfected with the recombinant genes. Furthermore, the soluble TCR is biologically active: it specifically inhibits antigen-dependent activation of the relevant T-cell clones and thus discriminates between proper and irrelevant peptides presented by major histocompatibility complex molecules.

Eosinophil-mediated diseases, such as allergic asthma, eosinophilic fasciitis, and certain hypersensitivity pulmonary disorders, are characterized by eosinophil infiltration and tissue injury. Mast cells and T cells often colocalize to these areas. Recent data suggest that mast cells can contribute to eosinophil-mediated inflammatory responses. Activation of mast cells can occur by antigen and immunoglobulin E (IgE) via the high-affinity receptor (FcepsilonRI) for IgE. The liberation of proteases, leukotrienes, lipid mediators, and histamine can contribute to tissue inflammation and allow recruitment of eosinophils to tissue. In addition, the synthesis and expression of a plethora of cytokines and chemokines (such as granulocyte-macrophage colony-stimulating factor [GM-CSF], interleukin-1 [IL-1], IL-3, IL-5, tumor necrosis factor-alpha [TNF-alpha], and the chemokines IL-8, regulated upon activation normal T cell expressed and secreted [RANTES], monocyte chemotactic protein-1 [MCP-1], and eotaxin) by mast cells can influence eosinophil biology. Stem cell factor (SCF)-c-kit, cytokine-cytokine receptor, and chemokine-chemokine receptor (CCR3) interactions leading to nuclear factor kappaB (NF-kappaB), mitogen-activated protein kinase (MAPK) expression, and other signaling pathways can modulate eosinophil function. Eosinophil hematopoiesis, activation, survival, and elaboration of mediators can all be regulated thus by mast cells in tissue. Moreover, because eosinophils can secrete SCF, eosinophils can regulate mast cell function in a paracrine manner. This two-way interaction between eosinophils and mast cells can pave the way for chronic inflammatory responses in a variety of human diseases. This review summarizes this pivotal interaction between human mast cells and eosinophils.

We have studied the relative contributions of monomeric (m-) and polymeric IgA (p-IgA) and of IgA1 and IgA2 to total serum IgA in healthy adults and patients with liver disease (LD) or with other diseases and high serum IgA. Serum concentration of total secretory component (SC) was also determined. In addition, fractional catabolic rates (FCR) and synthetic rates for both m- and p-IgA were measured in nine controls and nine cirrhotics. Our results support four main conclusions: (a) In healthy adults, intravascular p-IgA contributes to only 4-22% (mean 12%) of serum IgA, because its FCR and synthetic rate are approximately two times higher and four times smaller, respectively, than those of intravascular m-IgA. (b) in LD, biliary obstruction does not result in a significant increase in serum p-IgA unlike in rats and rabbits, indicating that in humans the SC-dependent biliary transport of p-IgA plays a much less significant role in selective removal of p-IgA from plasma than in rats and rabbits. (c) In contrast to biliary obstruction, parenchymal LD results in a significant and preferential increase in serum p-IgA, which in cirrhotics correlates with a selective reduction of the p-IgA-FCR. This supports a role for the human liver in selective removal of p-IgA from plasma, but another mechanism than the SC-dependent biliary transport should be considered. (d) Total SC, p-IgA, and IgA2 in serum are unlinked parameters, not necessarily reflecting mucosal events. A marked increase in serum SC occurs almost selectively in LD. Although a shift to IgA2 is suggested in Crohn's disease and alcoholic cirrhosis, a shift to IgA1 frequently associated to a shift to p-IgA occurs in chronic active LD, primary Sicca, and connective tissue diseases.

BACKGROUND

Japanese encephalitis virus (JEV), the mosquito-borne flavivirus, annually causes an estimated 35,000-50,000 encephalitis cases and 10,000-15,000 deaths in Asia, and there is no antiviral treatment. The role played by the immune response in determining the outcome of human infection with JEV is poorly understood, although, in animal models of flavivirus encephalitis, unregulated proinflammatory cytokine responses can be detrimental.

METHODS

We studied the innate, cellular, and humoral immune responses in 118 patients infected with JEV, of whom 13 (11%) died.

RESULTS

Levels of interferon (IFN)- alpha , the proinflammatory cytokine interleukin (IL)-6, and the chemokine IL-8 were all higher in the cerebrospinal fluid (CSF) of the nonsurvivors than of the survivors (P=.04, P=.006, and P=.04, respectively), as were both the IL-6 : IL-4 ratio in CSF (a marker of the balance of pro- and anti-inflammatory cytokines) and the level of the chemokine RANTES (regulated on activation, normally T cell expressed and secreted) in plasma (P=.03). In contrast, levels of immunoglobulin (Ig) M and IgG in CSF and of IgM in plasma were higher in the survivors (P=.035, P=.003, and P=.009, respectively). Levels of IFN- gamma and nitric oxide did not vary with outcome.

CONCLUSIONS

During JEV infection, elevated levels of proinflammatory cytokines and chemokines are associated with a poor outcome, but whether they are simply a correlate of severe disease or contribute to pathogenesis remains to be determined.

Acute renal insufficiency is a common problem, yet one that is frequently reversible with proper diagnosis and treatment. Although it has been argued that a renal biopsy is not needed for diagnosis in most cases of acute renal failure in the elderly, other studies have shown frequent disagreements between clinical and renal biopsy diagnoses in such cases. To investigate the causes of acute renal insufficiency in patients aged at least 60 years who underwent a renal biopsy and possible correlations between biopsy findings and renal survival, we first identified all native renal biopsy specimens from patients aged 60 years or older processed at The University of Chicago Medical Center (Chicago, IL) from 1991 through 1998 and reviewed the clinical records to determine the indication for the biopsy. We then reviewed again the records of those patients who underwent biopsy because of acute renal insufficiency, recorded the primary renal biopsy diagnosis in each of these cases, and obtained follow-up information for patients who underwent biopsy before July 1996. During the study period, 1,065 of 4,264 biopsy specimens (25.0%) received were obtained from patients aged 60 years or older, and acute renal insufficiency was the indication for biopsy in 259 of these patients (24.3%). The most frequent primary diagnoses on these latter biopsy specimens were pauci-immune crescentic glomerulonephritis (GN) with or without arteritis, 31.2% of biopsy specimens; acute interstitial nephritis, 18.6%; acute tubular necrosis (ATN) with nephrotic syndrome, 7.5%; atheroemboli, 7.1%; ATN alone, 6.7%; light chain cast nephropathy (LCCN), 5.9%; postinfectious GN, 5.5%; anti-glomerular basement membrane antibody nephritis, 4.0%; and immunoglobulin A (IgA) nephropathy and/or Henoch-Schönlein nephritis, 3.6%. Eight biopsy specimens (3.2%) showed only benign nephrosclerosis without an apparent cause of acute renal insufficiency, and another six specimens were inadequate. The renal biopsy diagnosis was in agreement with the prebiopsy clinical diagnosis (or differential diagnosis) in 107 of the 161 cases (67%) in which such information was provided. The distribution of diagnoses was similar in patients in the age groups of 60 to 69, 70 to 79, and 80 years or older, although younger age correlated significantly with improved renal and patient survival. The relative risk for progression to end-stage renal disease (ESRD) also increased according to diagnostic categories: LCCN (greatest risk)>> GN other than pauci-immune>> atheroemboli congruent with pauci-immune crescentic GN>> tubulointerstitial diseases other than LCCN (the latter category including ATN with nephrotic syndrome). Development of ESRD correlated significantly with decreased patient survival. In summary, renal biopsy in patients aged 60 years or older with acute renal insufficiency uncovered the cause in greater than 90% of the cases and provided clinically useful information with respect to expectation for renal survival and potential treatment options.

To determine the role of humoral mucosal immune response in protection against shigellosis, we have obtained a monoclonal dimeric immunoglobulin A (IgA) antibody specific for Shigella flexneri serotype 5a lipopolysaccharide (mIgA) and used a murine pulmonary infection model that mimics the lesions occurring in natural intestinal infection. Adult BALB/c mice challenged with 10(7) S. flexneri organisms developed a rapid inflammatory response characterized by polymorphonuclear cell infiltration around and within the bronchi and strong systemic interleukin 6 response. Implantation of hybridoma cells in the back of mice, resulting in the development of a myeloma tumor producing mIgA in the serum and subsequently secretory mIgA in local secretions, or direct intranasal administration of these antibodies, protected the animals against subsequent intranasal challenge with S. flexneri serotype 5a. Absence of histopathological lesion and significant decrease in bacterial load of the lungs and of systemic interleukin 6 response were the three major criteria of protection. This protection was shown to be serotype-specific and dependent on local concentration of mIgA. These data demonstrate that mucosal antibodies directed against a single polysaccharidic surface epitope of Shigella can protect against the disease.

Passively released or actively secreted elastase from neutrophils has been linked to the pathologic processes of a variety of inflammatory diseases, including idiopathic pulmonary fibrosis, rheumatoid arthritis, adult respiratory distress syndrome, and cystic fibrosis. The serine proteinase has a broad substrate specificity and may attack a number of host proteins outside of the neutrophil, including lung elastin and fibronectin. Such a proteolysis may change the normal surrounding tissue and the protein pattern of an inflammatory focus. Additionally, it acts as a potent secretagogue in minute amounts. The reason that neutrophil elastase is present in considerable concentrations outside of the neutrophil during chronic inflammation and that the major endogenous serine proteinase inhibitor for neutrophil elastase, alpha 1-proteinase inhibitor, is easily inactivated by proteolytic and oxidative attack is unclear. Released neutrophil elastase may also be involved in regulating chronic inflammation. In a feedback mechanism, neutrophil elastase inhibits neutrophil stimulation and concomitant elastase release by cleavage of immunoglobulins, complement components, and complement receptor type 1 on neutrophils. Besides a number of harmful effects of neutrophil elastase in inflammation, the latter mechanism, although considerably impairing phagocytosis, may be beneficial particularly in the light of persistent bacterial pathogens in the human lung affected by cystic fibrosis.

Through cognate B-cell-T-cell interactions and provision of cytokines, CD4+ T-cell antigen receptor (TCR) alpha beta+ T cells regulate immunoglobulin isotype synthesis. Murine IgG1 and IgE secretion is therefore substantially T-cell-dependent, whereas IgM and IgG3 secretion is not. Here we report that in the absence of alpha beta T cells, B cells expand, differentiate and secrete copious amounts of antibodies of 'T-dependent' isotypes. Moreover, the antibodies are reactive towards self-antigens, as in patients with systemic lupus erythematosus, so autoantibodies of 'T-dependent' type can develop without the help of CD4+ alpha beta T cells. This phenotype is not evident in mice or humans that are congenitally deficient in specific alpha beta T-cell functions, but bears comparison with B-cell hyperactivity and autoimmunity in transplant rejection and in immunodeficiencies such as AIDS.

The production and role of tumor necrosis factor alpha (TNF-alpha) in pneumococcal pneumonia were investigated in a mouse pneumonia model. When approximately 10(6) CFU of Streptococcus pneumoniae TUM19 were used to inoculate CBA/J mice intranasally, TNF-alpha levels in the lungs and serum began to increase from 1 and 3 days after infection, respectively, concomitantly with the increase in bacterial counts in the lungs. Anti-TNF-alpha antibody accelerated bacterial proliferation in the blood and the death of the mice. Although serum levels of immunoglobulin G antibody against the infecting bacteria were not affected by the anti-TNF-alpha antibody treatment, neutrophil counts in the blood were decreased by the treatment. These results suggest that TNF-alpha produced in the course of pneumococcal pneumonia prevents bacteremia by increasing the number of neutrophils in the blood.

Newborn mice suckled on dams immunized either orally or parenterally with primate rotavirus SA-11 were protected against diarrhea induced by SA-11 virus challenge. Experimental oral administration of milk from orally immunized dams protected suckling mice against challenge; protective activity was detected both in the anti-rotavirus immunoglobulin A (IgA) and IgG fractions, but IgA was more potent in vivo than IgG. Oral administration of milk from parentally immunized dams also protected suckling mice against challenge; in this case, protective activity was detected in the anti-rotavirus IgG fraction. In newborn mice foster-nursed by seronegative dams, circulating rotavirus-specific antibodies in high titer did not protect mice against oral SA-11 virus challenge. It appears that the most effective rotavirus vaccine will be that which induces an efficient production of antibodies active at the intestinal cell surface.

We have determined the complete nucleotides sequence (2168 bases) of the immunoglobulin mu gene cloned from newborn mouse DNA. The cloned 13kb fragment contained the entire constant region gene sequence that is interrupted by three intervening sequences at the junction of domains as previously shown in the gamma 1, gamma 2 b and alpha genes. The amino acid sequence predicted by the nucleotide sequence agrees with that of the mu chain secreted by a myeloma MOPC104E except for 8 residues out of 448 residues. The homologous domains of the mu, gamma 1 and gamma 2b genes are more similar to each other than the different domains of the mu genes are. The result implicates that the class of the immunoglobulin heavy chain genes diverged after the heavy chain genes established the multi-domain structure. The short intervening sequences of the mu and gamma genes are more conserved than the coding sequences except for the COOH-terminal domains. The results implicate that the nucleotide sequence of the intervening sequence is under selective pressure, possibly to maintain a secondary structure of the nuclear RNA to be spliced.

Francisella tularensis is a gram-negative intracellular bacterium that is considered to be a potential category A biological weapon due to its extreme virulence. Although vaccination with the attenuated live vaccine strain (LVS) of F. tularensis can protect against lethal challenge, use of inactivated or subunit forms as vaccine candidates for induction of protective antibody responses has not been fully evaluated. In the present study, we examined whether immune protection in the lung could be stimulated by intranasal administration of inactivated LVS together with interleukin-12 (IL-12) as an adjuvant. LVS was inactivated by heat, paraformaldehyde treatment, or exposure to UV, and inactivation of the preparations was confirmed by assessing bacterial growth and the survival of mice after direct inoculation. We found that mucosal vaccination with inactivated LVS provided 90 to 100% protection in mice after lethal intranasal challenge with 10(4) CFU of LVS, and this protection was dependent on inclusion of exogenous IL-12 during vaccine administration. Survival of vaccinated mice after live bacterial challenge was correlated with reduced bacterial burden, decreased pulmonary inflammation, increased serum antibody titers, and lower levels of gamma interferon (IFN-gamma), tumor necrosis factor alpha, and IL-6 in the lungs, livers, and spleens. Whereas NK cells were primarily responsible for the production of IFN-gamma in unvaccinated, challenged animals, vaccinated mice had increased levels of lung IFN-gamma+ CD4+ T cells after challenge. Significantly, mice genetically deficient in immunoglobulin A (IgA) expression were unable to survive lethal challenge after vaccination. These results are the first results to demonstrate that IgA-mediated protection against lethal respiratory tularemia occurs after mucosal vaccination with inactivated F. tularensis LVS.

Previous studies have shown that induction of autoimmune diabetes by adult thymectomy and split dose irradiation of PVG.RT1(u) rats can be prevented by their reconstitution with peripheral CD4(+)CD45RC-TCR-alpha/beta+RT6(+) cells and CD4(+)CD8(-) thymocytes from normal syngeneic donors. These data provide evidence for the role of regulatory T cells in the prevention of a tissue-specific autoimmune disease but the mode of action of these cells has not been reported previously. In this study, autoimmune thyroiditis was induced in PVG.RT1(c) rats using a similar protocol of thymectomy and irradiation. Although a cell-mediated mechanism has been implicated in the pathogenesis of diabetes in PVG.RT1(u) rats, development of thyroiditis is independent of CD8(+) T cells and is characterized by high titers of immunoglobulin (Ig)G1 antithyroglobulin antibodies, indicating a major humoral component in the pathogenesis of disease. As with autoimmune diabetes in PVG. RT1(u) rats, development of thyroiditis was prevented by the transfer of CD4(+)CD45RC- and CD4(+)CD8(-) thymocytes from normal donors but not by CD4(+)CD45RC+ peripheral T cells. We now show that transforming growth factor (TGF)-beta and interleukin (IL)-4 both play essential roles in the mechanism of this protection since administration of monoclonal antibodies that block the biological activity of either of these cytokines abrogates the protective effect of the donor cells in the recipient rats. The prevention of both diabetes and thyroiditis by CD4(+)CD45RC- peripheral cells and CD4(+)CD8(-) thymocytes therefore does not support the view that the mechanism of regulation involves a switch from a T helper cell type 1 (Th1) to a Th2-like response, but rather relies upon a specific suppression of the autoimmune responses involving TGF-beta and IL-4. The observation that the same two cytokines were implicated in the protective mechanism, whether thymocytes or peripheral cells were used to prevent autoimmunity, strongly suggests that the regulatory cells from both sources act in the same way and that the thymocytes are programmed in the periphery for their protective role. The implications of this result with respect to immunological homeostasis are discussed.

Immunoglobulin class switch involves a unique recombination event that takes place at the switch (S) region which is located 5' to each constant region (C) gene of the heavy (H) chain. For example, differentiation of the B lymphocyte from a mu-chain producer to an epsilon-chain producer is mediated by the switch recombination between the S mu and S epsilon regions. In order to elucidate the molecular mechanism for the switch recombination, we have determined nucleotide sequences surrounding the class switch recombination sites of the C epsilon and C gamma 3 genes and those in the 5' flanking regions of the C gamma 2a and C delta genes. The results indicate that the 5' flanking regions of all the CH genes except for the C delta gene contain the S regions which comprise tandem repetition of short unit sequences in agreement with the previous analyses of the S gamma 1, S gamma 2b, S mu, and S alpha regions. Comparison of the nucleotide sequences of all the S regions revealed that length as well as nucleotide sequences of the S regions vary among different classes of the CH gene, but they share short common sequences, (G)AGCT and TGGG(G). The nucleotide sequence of the S mu region is homologous to those of the other S regions in the decreasing order of the S epsilon, S alpha, S gamma 3, and (S gamma 1, S gamma 2b, s gamma 2a) regions. We have compared the nucleotide sequences immediately adjacent to the recombination sites of seven rearranged genes and have always fund tetranucleotides TGAG and/or TGGG, except for one case. Such tetranucleotides may constitute a part of the recognition sequence of a putative recombinase. These results provide further support for our previous proposal that the switch recombination may be facilitated by short common sequences dispersed in all the S regions.

Swine influenza viruses (SIV) naturally infect pigs and can be transmitted to humans. In the pig, genetic reassortment to create novel influenza subtypes by mixing avian, human, and swine influenza viruses is possible. An SIV vaccine inducing cross-protective immunity between different subtypes and strains circulating in pigs is highly desirable. Previously, we have shown that an H3N2 SIV (A/swine/Texas/4199-2/98 [TX98]) containing a deleted NS1 gene expressing a truncated NS1 protein of 126 amino acids, NS1black triangle126, was attenuated in swine. In this study, 4-week-old pigs were vaccinated with the TX98 NS1black triangle126 modified live virus (MLV). Ten days after boosting, pigs were challenged with wild-type homologous H3N2 or heterosubtypic H1N1 SIV and sacrificed 5 days later. The MLV was highly attenuated and completely protected against challenge with the homologous virus. Vaccinated pigs challenged with the heterosubtypic H1N1 virus demonstrated macroscopic lung lesions similar to those of the unvaccinated H1N1 control pigs. Remarkably, vaccinated pigs challenged with the H1N1 SIV had significantly lower microscopic lung lesions and less virus shedding from the respiratory tract than did unvaccinated, H1N1-challenged pigs. All vaccinated pigs developed significant levels of hemagglutination inhibition and enzyme-linked immunosorbent assay titers in serum and mucosal immunoglobulin A antibodies against H3N2 SIV antigens. Vaccinated pigs were seronegative for NS1, indicating the potential use of the TX98 NS1black triangle126 MLV as a vaccine to differentiate infected from vaccinated animals.

Vaccinia virus contains a gene, termed SPI-2 or B13R, that is closely related in its sequence to a potent inhibitor of apoptosis from cowpox virus (crmA). Infection by vaccinia virus protects HeLa cells against apoptosis that is induced by an immunoglobulin M antibody against the fas receptor or by tumor necrosis factor alpha. This effect is profoundly reduced when the SPI-2 gene is deleted. The SPI-2 gene, when transiently expressed in these cells, can also protect against apoptosis mediated by these agents. Given the similarity to crmA, it seems likely that SPI-2 functions in an analogous fashion, inhibiting the activity of ICE protease family members and blocking the onset of apoptosis.

OBJECTIVE

To determine the natural history of immunoglobulin (Ig) A nephropathy among patients who presented with hematuria and minimal proteinuria, and factors associated with the development of adverse clinical events, such as proteinuria.

METHODS

In Hong Kong, all patients who present with isolated hematuria are referred for renal biopsy after urologic diseases are ruled out. We reviewed the clinical course of 72 consecutive patients with histologically confirmed IgA nephropathy who presented with hematuria and minimal proteinuria (0.4 g/day or less). All patients were normotensive and had normal renal function at presentation. Adverse events were defined as proteinuria greater than 1 g per day, hypertension, or impaired renal function (serum creatinine level 120 micromol/L or estimated creatinine clearance < 70 mL per minute).

RESULTS

The mean (+/- SD) age at presentation was 27 +/- 8 years; 56 (78%) were female. Nine patients (13%) had grade 2 histologic lesions. During a median follow-up of 7 years, 32 patients (44%) developed adverse events: 24 (33%) developed proteinuria of 1 g per day or more, 19 (26%) became hypertensive, and 5 (7%) developed impaired renal function. Another 30 patients (42%) had persistently abnormal urinalysis examinations. Only 10 patients (14%) had complete resolution of hematuria. The median time for progression from proteinuria >> l g/day) to renal impairment was 84 months (range 56 to 132). In a multivariate analysis, age at presentation (relative risk [RR] per 10 years of age = 2.0; 95% confidence interval [CI], 1.2 to 3.4) and histologic grade (grade 2 versus grade 1, RR = 4.5; 95% CI, 1.7 to 12) were independent predictors of developing an adverse event.

CONCLUSIONS

IgA nephropathy that presents with hematuria and minimal proteinuria is usually a progressive disease. Life-long follow-up with regular monitoring of blood pressure and proteinuria is recommended.