Systemic lupus erythematosus aggravates atherosclerosis by promoting IgG deposition and inflammatory cell imbalance.

BACKGROUND: Systemic lupus erythematosus (SLE) patients experience a premature and more severe presentation of coronary artery disease. The underlying mechanisms of accelerated coronary artery disease in SLE patients remain to be elucidated. METHODS: By using atherosclerosis combining a SLE murine model, we proved that the onset of SLE aggravates atherosclerosis. Although the onset of SLE reduced blood lipids slightly, immune deviation contributed to aggravated atherosclerosis in lupus mice. Lupus atheroma were characterized by inflammatory cell infiltration, such as gathered dendritic cells, macrophages, and IgG deposition. RESULTS: Decreased lymphocytes and magnified dendritic cells in the spleen were also observed in lupus mice. Hydroxychloroquine prevented atherosclerosis progression mainly by reversing immune status abnormality caused by SLE. Serum interferon alfa levels were not changed in lupus mice. CONCLUSION: These findings strongly suggested that anti-inflammatory therapies and hydroxychloroquine provide a new possible strategy for treating SLE patients with atherosclerosis.

Intracellular and extracellular serpins modulate lung disease.

An imbalance between peptidases and their inhibitors leads to pulmonary disease. Imbalances occur in the adult and the neonate at risk for a specific set of lung pathologies. Serpins (serine peptidase inhibitors) make up the major source of antipeptidase activity in the lung. The purpose of this review is to describe the serpin mechanism of inhibition, their roles in the normal and pathological lung and their potential as therapeutic agents.

Rituximab treatment of pulmonary arterial hypertension associated with systemic lupus erythematosus: a case report.

Pulmonary hypertension is a common but underdiagnosed complication of systemic lupus erythematosus, which can be associated with significant morbidity and early mortality. Although often associated with anti-phospholipid antibodies, the etiology remains poorly understood. In case reports and small open trials, the anti-CD20, B-cell targeted therapeutic antibody, rituximab, has been reported to provide benefits for systemic lupus erythematosus patients with glomerulonephritis, anti-phospholipid antibody syndrome, vasculitis, arthritis, and refractory skin disease. However, the outcome of rituximab treatment of pulmonary arterial hypertension associated with systemic lupus erythematosus has not been described. We, therefore, present a case of a young systemic lupus erythematosus patient with early onset of pulmonary arterial hypertension during the disease course, refractory to multiple treatment modalities, who had significant improvement with rituximab therapy.

Genetic imprinting of autoantibody repertoires in systemic lupus erythematosus patients.

Systemic lupus erythematosus (SLE) is an autoimmune disease distinguished by great heterogeneity in clinical manifestations and autoantibody expression. While only a handful of autoantibody specificities have proved useful for clinical diagnosis, to characterize complex lupus-associated autoantibody profiles more fully we have applied proteome microarray technology. Our multiplex microarrays included control ligands and 65-autoantigens, which represent diverse nuclear and cytoplasmic antigens recognized by disease-associated and natural autoantibodies. From longitudinal surveys of unrelated SLE patients, we found that autoantibody profile patterns can be patient-specific and highly stable overtime. From profiles of 38 SLE patients that included 14 sets of SLE twins, autoantibodies to the phospholipid neo-determinants, malondialdehyde (MDA) and phosphorylcholine (PC), which are exposed on apoptotic but not healthy cells, were among the most prevalent and highly expressed. We also found that immunoglobulin M (IgM) reactivity to MDA and PC ligands had significant direct correlations with DNA-containing antigens, while such a general relationship was not found with a panel of RNA-related antigens, or for IgG-autoantibodies. Significantly, hierarchical analysis revealed co-distribution/clustering of the IgM autoantibody repertoire patterns for six of 14 twin sets, and such patterns were even more common (10 of 14) for IgG autoantibody profiles. Our findings highlight the potentially distinct roles of IgM and IgG autoantibodies, as we postulate that the direct correlations for IgM autoantibodies to DNA antigens with apoptosis-related determinants may be due to co-expression arising from common pro-homeostatic protective roles. In contrast, the sharing of IgG autoantibody fingerprints by monozygotic twins suggests that lupus IgG autoantibodies can arise in predisposed individuals in genetically determined patterns.

Characterization of monoclonal antibodies directed against squamous cell carcinoma antigens: report of the TD-10 Workshop.

Thirteen monoclonal antibodies directed against squamous cell carcinoma antigens (SCCA1 and SCCA2) were obtained from five international collaborating laboratories participating in the ISOBM TD-10 Workshop. Native and recombinant forms of SCCA were used in a wide variety of approaches to determine the reactivity and specificity of these antibodies. Based on reactivity, the antibodies could be divided into three groups: the SCCA1-reactive group containing those that reacted only with recombinant SCCA1 (rSCCA1) and native SCCA1 (nSCCA1) antigens, the SCCA2-reactive group containing those that reacted only with recombinant SCCA2 (rSCCA2), and the pan-reactive group containing those antibodies that reacted with rSCCA1, nSCCA1, and rSCCA2. Binding to radioiodinated rSCCA1 showed that all reactive antibodies were of a high affinity (K(d) <2 x 10(-9) mol/l). Binding to labelled rSCCA2 demonstrated that five antibodies were of a high affinity (K(d) <2 x 10(-9) mol/l). Antibody reactivity on Western blots was tested with nonreduced and reduced native and recombinant SCCA1 and SCCA2. In general, these findings showed that reduction had little effect on binding to SCCA1, but often a strong effect on the binding to SCCA2. Binding of antibodies to rSCCA1 and rSCCA2 in complexes with cathepsin L and G, respectively, was used to assist in the localization of epitope regions in enzyme-complexed SCCA. Cross-inhibition experiments showed that SCCA1-reactive antibodies represent two different epitope groups, and this is supported by their ability to make SCCA1-specific assays by combining antibodies from the two epitope groups. The SCCA2-reactive group represents two related antibodies and one unique as seen in cross-inhibition, but they do not form successful assay combinations. Classification of the pan-reactive antibodies is more difficult, as some epitope groups differ when results from rSCCA1 are compared with rSCCA2 as the target. However, two antibodies are outstanding, SCC107 and SCC113, as they are high-affinity antibodies which react equally well with free and protease complexes of SCCA1 and SCCA2. The precise location of epitopes was further studied using sequential overlapping peptides and homology modelling. The findings from this workshop strongly indicate that the recombinant antigens (rSCCA1 and rSCCA2) are very similar in epitope structure to the native counterparts in saliva, and squamous epithelium from normal and cancer tissues. Therefore, it is reasonable to conclude that the specificities found are reliable and have application for antibody measurement of all forms of squamous cell carcinoma in serum except SCCA2 in complex with its protease.

Human clade B serpins (ov-serpins) belong to a cohort of evolutionarily dispersed intracellular proteinase inhibitor clades that protect cells from promiscuous proteolysis.

Serpins are unique among the various types of active site proteinase inhibitors because they covalently trap their targets by undergoing an irreversible conformational rearrangement. Members of the serpin superfamily are present in the three major domains of life (Bacteria, Archaea and Eukarya) as well as several eukaryotic viruses. The human genome encodes for at least 35 members that segregate evolutionarily into nine (A-I) distinct clades. Most of the human serpins are secreted and circulate in the bloodstream where they reside at critical checkpoints intersecting self-perpetuating proteolytic cascades such as those of the clotting, thrombolytic and complement systems. Unlike these circulating serpins, the clade B serpins (ov-serpins) lack signal peptides and reside primarily within cells. Most of the human clade B serpins inhibit serine and/or papain-like cysteine proteinases and protect cells from exogenous and endogenous proteinase-mediated injury. Moreover, as sequencing projects expand to the genomes of other species, it has become apparent that intracellular serpins belonging to distinct phylogenic clades are also present in the three major domains of life. As some of these serpins also guard cells against the deleterious effects of promiscuous proteolytic activity, we propose that this cytoprotective function, along with similarities in structure are common features of a cohort of intracellular serpin clades from a wide variety of species.

Preliminary explorations of the effects of prior trauma and loss on risk for psychiatric disorders in recently widowed people.

BACKGROUND: This study compared the relative influence of childhood and adulthood adversities on current diagnoses of Major Depressive Episode (MDE), Post-Traumatic Stress Disorder (PTSD) and Traumatic Grief (TG) among recently widowed older adults. METHOD: Eighty-five widowed persons were interviewed at a median of 4 months after their loss. The logistic regression procedure was used to estimate the effects of three childhood adversities (parental death, abuse, death of a sibling) and three prior adulthood adversities (death of a child, non-bereavement traumatic event, death of a sibling) on current diagnoses of MDE, PTSD and TG. RESULTS: Adversities occurring in childhood (abuse and death of a parent) were significantly associated with TG and, secondarily, MDE, while adversities occurring in adulthood (non-bereavement traumatic event and death of a child) were only significantly associated with PTSD. The tendency of childhood adversities to predict TG and adult adversities to predict PTSD remained significant even after the clustering of adversities and comorbidity among psychiatric disorders were taken into account. CONCLUSIONS: The results suggest that there is a vulnerability to TG rooted in childhood experiences explicitly, with more recent traumas having a stronger influence on PTSD secondary to widowhood. The distinctive etiological risks for bereavement-related PTSD, MDE, and TG suggest that therapeutic approaches should be tailored to the particular syndrome(s) present.

Regulation of inherently autoreactive VH4-34 B cells in the maintenance of human B cell tolerance.

The study of human B cell tolerance has been hampered by difficulties in identifying a sizable population of autoreactive B lymphocytes whose fate could be readily determined. Hypothesizing that B cells expressing intrinsically autoreactive antibodies encoded by the VH4-34 heavy chain gene (VH4-34 cells) represent such a population, we tracked VH4-34 cells in healthy individuals. Here, we show that naive VH4-34 cells are positively selected and mostly restricted to the follicular mantle zone. Subsequently, these cells are largely excluded from the germinal centers and underrepresented in the memory compartment. In healthy donors but not in patients with systemic lupus erythematosus (SLE), these cells are prevented from differentiating into antibody-producing plasma cells. This blockade can be overcome ex vivo using cultures of naive and memory VH4-34 cells in the presence of CD70, IL-2, and IL-10. VH4-34 cells may therefore represent an experimentally useful surrogate for autoantibody transgenes and should prove valuable in studying human B cell tolerance in a physiological, polyclonal environment. Our initial results suggest that both positive and negative selection processes participate in the maintenance of tolerance in autoreactive human B cells at multiple checkpoints throughout B cell differentiation and that at least some censoring mechanisms are faulty in SLE.

SERPINB12 is a novel member of the human ov-serpin family that is widely expressed and inhibits trypsin-like serine proteinases.

Members of the human serpin family regulate a diverse array of serine and cysteine proteinases associated with essential biological processes such as fibrinolysis, coagulation, inflammation, cell mobility, cellular differentiation, and apoptosis. Most serpins are secreted and attain physiologic concentrations in the blood and extracellular fluids. However, a subset of the serpin superfamily, the ov-serpins, also resides intracellularly. Using high throughput genomic sequence, we identified a novel member of the human ov-serpin gene family, SERPINB12. The gene mapped to the ov-serpin cluster at 18q21 and resided between SERPINB5 (maspin) and SERPINB13 (headpin). The presence of SERPINB12 in silico was confirmed by cDNA cloning. Expression studies showed that SERPINB12 was expressed in many tissues, including brain, bone marrow, lymph node, heart, lung, liver, pancreas, testis, ovary, and intestines. Based on the presence of Arg and Ser at the reactive center of the RSL, SERPINB12 appeared to be an inhibitor of trypsin-like serine proteinases. This hypothesis was confirmed because recombinant SERPINB12 inhibited human trypsin and plasmin but not thrombin, coagulation factor Xa, or urokinase-type plasminogen activator. The second-order rate constants for the inhibitory reactions were 2.5 +/- 1.6 x 10(5) and 1.6 +/- 0.2 x 10(4) M(-1) S(-1), respectively. These data show that SERPINB12 encodes for a new functional member of the human ov-serpin family.

A randomized trial of stenting with or without balloon predilation for the treatment of coronary artery disease.

BACKGROUND: Stent placement has historically been preceded by predilation of the target lesion with percutaneous transluminal coronary angioplasty. Direct stent implantation, without predilation, has the potential to have a favorable impact on procedure cost by reducing the number of devices used, contrast administered, and procedure time. METHODS AND RESULTS: We conducted a prospective randomized trial to compare the economic outcome of stenting with or without predilation. Inclusion criteria included intention to treat a single lesion with a coronary stent in a vessel with a reference diameter >2.4 mm. Exclusion criteria included total occlusions, culprit lesion within a saphenous vein graft, lesion length >25 mm, patients within 48 hours of an acute myocardial infarction, and patients unable to be treated with aspirin and clopidogrel. From September 1999 to March 2000, 77 patients were randomized to direct stent implantation (n = 37) or balloon-facilitated stenting (n = 40). Stent placement was successful in all patients. Crossover to predilation was required in 2 patients in the direct stent group because of inability to deliver the stent. Compared with balloon-facilitated stenting, direct stenting used fewer catheter devices (1.4 +/- 0.7 vs 2.5 +/- 0.8, P <.001), less contrast (92.7 +/- 43.1 mL vs 117.4 +/- 61.0 mL, P =.04), and less fluoroscopy time (7.5 +/- 3.9 minutes vs 11.6 +/- 8.3 minutes, P =.006). No difference in procedural complications or predischarge outcome was found. No difference in major adverse cardiovascular events was found at 6-month follow-up. CONCLUSION: Direct stenting is a safe and successful procedure that reduces the number of devices used, fluoroscopy time, and contrast administration.

Human-derived anti-oxidized LDL autoantibody blocks uptake of oxidized LDL by macrophages and localizes to atherosclerotic lesions in vivo.

Autoantibodies to oxidation-specific epitopes of low density lipoprotein (LDL), such as malondialdehyde-modified LDL (MDA-LDL), occur in plasma and atherosclerotic lesions of humans and animals. Plasma titers of such antibodies are correlated with atherosclerosis in murine models, and several such autoantibodies have been cloned. However, human-derived monoclonal antibodies to epitopes of oxidized LDL (OxLDL) have not yet been reported. We constructed a phage display antibody library from a patient with high plasma anti-MDA-LDL titers and isolated 3 monoclonal IgG Fab antibodies, which specifically bound to MDA-LDL. One of these, IK17, also bound to intact OxLDL as well as to its lipid and protein moieties but not to those of native LDL. IK17 inhibited the uptake of OxLDL by macrophages and also bound to apoptotic cells and inhibited their phagocytosis by macrophages. IK17 strongly immunostained necrotic cores of human and rabbit atherosclerotic lesions. When (125)I-IK17 was injected intravenously into LDL receptor-deficient mice, its specific uptake was greatly enriched in atherosclerotic plaques versus normal aortic tissue. Human autoantibodies to OxLDL have important biological properties that could influence the natural course of atherogenesis.

Adoptive transfer of a superantigen-induced hole in the repertoire of natural IgM-secreting cells.

We have recently evaluated the host response to the bacterial toxin, protein A from Staphylococcus aureus (SpA), which has the capacity to interact with B-cell antigen receptors encoded by V(H) clan III genes via a conserved variable region framework surface. In these studies, intraperitoneal instillation of SpA induced a persistent T-cell independent loss of a large supraclonal set of susceptible lymphocytes, which includes clan III/V(H) S107 family-expressing B-1 cells and their antibody products. To determine whether these long-term effects could represent the influence of residual in vivo deposited superantigen, we have now performed adoptive transfer of peritoneal B cells from superantigen- and control-treated donors. These studies demonstrated that mice that received cells from SpA-treated donors also exhibited the same induced supraclonal hole in the expressed repertoire of natural IgM-secreting cells due to supraclonal deletion. These studies clarify the cellular mechanisms responsible for B-cell superantigen-induced modification of the repertoires of in vivo polyclonal B-cell populations.

Undergraduate environmental health education: preparing for the future.

Attendees indicated that the workshop was beneficial and that the opportunity to communicate with faculty representing 23 programs accredited by EHAC and nine programs not accredited by EHAC was extremely useful. There was general agreement on a number of points: There is a need for undergraduate environmental health programs to collaborate on matters related to distance learning. Topics related to women, gender, diversity, ethics, and international environmental health should be incorporated into the environmental health curriculum. There are no major problems with current EHAC curricular guidelines, but the guidelines should be evaluated on a regular basis. Field experience or internship is an essential component in the academic preparation of undergraduate environmental health students. There is a significant need for increased funding for undergraduate environmental health programs. There is a need to increase the visibility and recognition of environmental health programs. There is a need to solidify ties with traditional partners and to establish new linkages at the local, regional, and national levels in the government, community, and private sector. It is essential that undergraduate faculty communicate with each other on matters relating to the preparation of environment health practitioners. There is a need for an association of undergraduate academic programs to provide leadership and a focal point for identification and resolution of issues common to all. The establishment of an association was viewed as the most practical and effective way to address these issues and to pursue related opportunities.

SCCA1 expression in T-lymphocytes peripheral to cancer cells is associated with the elevation of serum SCC antigen in squamous cell carcinoma of the tongue.

Squamous cell carcinoma (SCC) antigen has been used for the management of SCC arising in various cites including head and neck region. However, the true mechanism of the elevation of this protein in the serum of patients with SCC is still unknown. SCC antigen belongs to the superfamily of serine protease inhibitors. Recently, molecular studies show that serum SCC antigen is transcribed by two nearly identical genes (SCCA1 and SCCA2), and is mainly produced by SCCA1. The objective of this study is to clarify the mechanism of the elevation of SCC antigen in oral tongue SCC patients and to identify cells histologically, which are responsible for serum SCC antigen production. In this study, we examined SCCA1 expression in a series of four head and neck SCC (HNSCC) cell lines, and found that all expressed equal to low SCCA1 protein as compared with the normal human oral keratinocyte. Using the double immunohistochemical technique to examine the expression pattern of SCCA1 in 86 cases of oral tongue squamous cell carcinoma, SCCA1 immunostaining was observed in the cytoplasm of cancer cells and T-lymphocytes peripheral to cancer cells. We also compared the clinicopathological features including serum SCC antigen level of the oral tongue SCC cases with the immunohistochemical SCCA1 expression pattern, and found that elevated serum SCC antigen level was significantly correlated with SCCA1 expression not in cancer cells, but in T-lymphocytes peripheral to cancer cells. These results suggest that T-lymphocytes peripheral to cancer cells may be responsible for serum SCC antigen production in HNSCC patients.

Suppression of a squamous cell carcinoma (SCC)-related serpin, SCC antigen, inhibits tumor growth with increased intratumor infiltration of natural killer cells.

Squamous cell carcinoma (SCC) antigen (SCCA), a member of the ovalbumin serine proteinase inhibitor family, serves as a circulating marker of squamous cell carcinoma (SC). One of the SCCAs, SCCA1, has been suggested to play a role in the attenuation of apoptosis in vitro and in the augmentation of tumor growth in vivo. In the present study, the infection of a SCC cell line (SKG IIIa) with recombinant retrovirus that expressed the antisense SCCA mRNA suppressed expression of SCCA in vitro. Local administration of this retrovirus into tumors by inoculation in nude mice suppressed tumor growth. Treatment of tumor tissue in vivo is also associated with increased numbers of apoptotic tumor cells and large mononuclear cells in the tumor. To test the possible role of SCCA in the infiltration of large mononuclear cells, we analyzed the effect of SCCA1 on migration of natural killer (NK) cells induced by monocyte-chemoattractant protein-1 in vitro. SCCA1 suppressed migration of NK cells completely, and this inhibitory effect was lost by mutation of the reactive site loop of SCCA1. These results suggest that antisense SCCA may suppress the growth of SCC in vivo not only by the augmentation of intracellular apoptosis but also by the increased infiltration of NK cells into the tumor.

Building community capacity in public health: the role of action-oriented partnerships.

Public health practice increasingly is concerned with the capacity and performance of communities to identify, implement, strengthen, and sustain collective efforts to improve health. The authors developed ways to assist local Turning Point partnerships to improve their community public health system as a secondary outcome of their work on the expressed needs of the community. Using focus groups, meeting minutes, attendance records, and meeting observation, the authors fed information back to the partnerships on systems change. A public health systems improvement plan supportive of local partnerships' work on specific health issues was funded and the collaborative research agenda was further refined.

Building larger YACs by recombination.

Despite the relatively large cloning capacity of YACs, many genomic regions or individual genes are not cloned intact, but are represented as a collection of overlapping clones or contigs. Fortunately, the relatively high frequency and fidelity of homologous recombination in Saccharomyces cerevisiae can be used to reconstruct intact genes within a single clone by splicing together overlapping DNA segments. This unit describes two protocols for carrying out such homologous recombination; one relies on the meiotic phase of the yeast cycle, while the other utilizes the mitotic phase of the yeast life cycle. Despite the relatively large cloning capacity of YACs, many genomic regions or individual genes are not cloned intact.

Neo-self antigens and the expansion of B-1 cells: lessons from atherosclerosis-prone mice.

The pathogenesis of atherosclerosis involves an inflammatory process that is modulated by the immune system, and within these complex responses we have discerned a possible role for an archetypic B-1 clone. We speculate that due to their immunogenicity and in vivo distribution the "neo"-self determinants created in oxidatively modified LDL are highly stimulatory for certain B-1 cell clones. These neo-self determinants, which can be created chemically, by somatic processes, may in fact represent the molecular analogues of somatic maturation, or even aging. These changes, including those on non-protein antigens induced by oxidative metabolism, amongst others, create neo-determinants against which the host no doubt can not develop rigorous B-cell tolerance. The onset of expression of these oxidative neo-determinants relatively late in development may well serve a useful function for the highly evolved mammalian immune system, as targeting by evolutionarily selected B-1 clones may facilitate the amplification of other useful antibody-mediated physiologic functions. As in the case of the T15 clone, these antibodies may aid in protection against common microbial pathogens. Hence we postulate that during the evolution of the adaptive immune system the neo-self antigenic milieu may have been exploited for the natural selection of primordial clonal specificities. The T15 B-1 clone may then illustrate a common paradigm in which there has been natural selection based on utility for the defense of the individual from environmental threats, as well as for possible "housekeeping" role(s) and the maintenance of cellular homeostasis.