Monoclonal anti-tumor necrosis factor antibody renders non-obese diabetic mice hypersensitive to irradiation and enhances insulitis development.

In attempt to evaluate biological roles of tumor necrosis factor (TNF), we studied the effects of anti-TNF mAb in non-obese diabetic (NOD) mice. Anti-murine TNF mAb rendered NOD mice hypersensitive to the lethal effects of radiation and prevented the reconstitution of lethally irradiated mice with adoptively transferred lymphocytes. While TNF-alpha reduced the incidence of diabetes development in the adoptive transfer system even when given 6 days post-transfer, mAb to TNF could not reduce or increase the incidence of diabetes compared to control mice. Administration of TNF-alpha for 4 or 8 weeks significantly reduced the incidence of spontaneous insulitis in NOD mice, while anti-TNF mAb given for 8 weeks increased the incidence of insulitis significantly.     

Transient hypogonadotrophic hypogonadism in males with acute toxoplasmosis: suppressive effect of interleukin-1 beta on the secretion of GnRH

BACKGROUND: In September 2002, an outbreak of toxoplasmosis was noted in a male boarding high school on the Aegean coast of Turkey. We have focused our efforts to investigate the sex hormones in this population. METHODS: Blood samples were collected from 40 male patients, 17-18 years old, who also had positive titres of antibody to Toxoplasma gondii. Serum FSH, LH, free testosterone (FT), total testosterone (TT), interferon-gamma (IFN-gamma), interleukin-1beta (IL-1beta) and macrophage-inflammatory protein-1alpha (MIP-1alpha) concentrations were measured in all patients and 20 control subjects. Initially, the patients were divided on the basis of the levels of sex hormones into the following groups: patients who had normal sex hormone levels (n = 31) as group A and patients with low sex hormone levels (n = 9) as group B. RESULTS: IL-1beta levels were found to be higher in group B patients than group A. The levels of IL-1beta correlated significantly in a negative manner with FSH, LH, FT and TT in all patients with acute toxoplasmosis (n = 40). CONCLUSIONS: Acute toxoplasma infection may cause temporary hypogonadotrophic gonadal insufficiency regardless of the course of the disease.     

Maternal serum levels of interferon-gamma and interleukin-2 soluble receptor-alpha predict the outcome of early IVF pregnancies

BACKGROUND: Elevated maternal serum levels of interleukin-2 soluble receptor-alpha (IL-2 sRalpha), tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) have been associated with pregnancy loss. The aim of our study was to evaluate the predictive value of these cytokines in the outcome of early IVF pregnancies. METHODS: One hundred and fifty-nine consecutive IVF patients who were subsequently diagnosed to have a biochemical pregnancy (n = 23), a first-trimester miscarriage (n = 19) or a normal term delivery (n = 117) were included in this study. Serum was collected from the initial pregnancy test, 11 days after a day 3 embryo transfer, and all samples were analysed for IL-2 sRalpha, TNF-alpha and IFN-gamma by commercially available enzyme-linked immunosorbent assay (ELISA) kits. RESULTS: IL-2 sRalpha levels were significantly higher in patients with an early pregnancy loss compared with patients with a normal term delivery (849.5 +/- 69.6 versus 693.5 +/- 31.2 pg/ml, P = 0.02), and a cut-off point of IL-2 sRalpha >1000 pg/ml predicted a poor pregnancy outcome (44.4 versus 22.7% pregnancy loss, IL-2 sRalpha >or=1000 versus IL-2 sRalpha <1000 pg/ml; P = 0.02). IFN-gamma-positive patients had twice the risk for poor IVF pregnancy outcome compared with IFN-gamma-negative subjects (40.8 versus 20.0%, respectively; P < 0.02), including a significantly lower implantation rate (37.6 +/- 0.05 versus 50.0 +/- 0.03%, respectively; P = 0.02). There was no difference in pregnancy outcome based upon serum levels, or the ability to detect the presence of TNF-alpha. No differences in levels of these cytokines were found based on the aetiology of the patients' infertility. CONCLUSIONS: Elevated maternal serum levels of IL-2 sRalpha and IFN-gamma as early as 11 days after embryo transfer are associated with poor IVF pregnancy outcome.     

Activation of Vgamma9Vdelta2 T cells by non-peptidic antigens induces the inhibition of subgenomic HCV replication

Hepatitis C virus (HCV) has evolved complex strategies to evade host immune responses and establish chronic infection. Since human Vgamma9Vdelta2 T lymphocytes play a critical role in the immune response against viruses, we analyzed their antiviral functions on Huh7 hepatoma cells carrying the subgenomic HCV replicon (Rep60 cells). In a transwell culture system, Rep60 cells were co-cultured with either PBMCs or highly purified gammadelta T cells stimulated by non-peptidic antigens. Vgamma9Vdelta2 T cell activation was associated with a dramatic reduction of HCV RNA levels. Neutralizing antibodies targeting IFN-gamma revealed a critical role for this cytokine in the inhibition of HCV replication. Interestingly, drugs already in clinical use, such as Phosphostim and Zoledronate, known to activate gammadelta T cells, were shown to induce the inhibition of HCV replication mediated by Vgamma9Vdelta2 T cells of HCV patients. Our data suggest that the therapeutic activation of Vgamma9Vdelta2 T lymphocytes may represent an additional strategy to inhibit HCV replication and to restore a Th1-oriented immune response in HCV-infected patients.     

Expression of a fetal gamma delta T-cell receptor in adult mice triggers a non-MHC-linked form of selective depletion.

Day 14 fetal thymocytes and adult dendritic epidermal T cells (dEC) of all mouse strains express a characteristic non-polymorphic gamma delta T-cell receptor which is rarely found in the adult thymus or lymph nodes. We have made transgenic mice expressing this particular set of receptors on T cells in C3H and C57BL/6 mice. In adult mice of the latter strain, a dramatic depletion of transgene expressing T cells occurs and this effect is primarily mediated by thymic radiosensitive cells. The depletion is genetically dominant but not MHC-linked with major factor(s) mapping to chromosome 18. Taken together, our results show that strain-specific developmental changes in the thymic environment may play a role in shaping the gamma delta TCR repertoire.     

Changes in the expression of macrophage antigens associated with activation for tumor cell killing

Cultured murine bone marrow macrophages (BMM phi) can be induced to kill tumor cells in vitro by combined treatment with a priming (IFN-gamma) and a triggering (LPS) agent. We have examined the expression of cellular antigens accompanying this activation by Western blot analysis with rabbit antisera raised against unstimulated and fully activated BMM phi and assayed the effect of these antisera on macrophage-mediated tumor cytotoxicity. Killing of Yac-1 target cells was slightly enhanced by antiserum against unstimulated BMM phi but inhibited 54% by antiserum against activated BMM phi. The following changes in antigen expression are shown to be associated with discrete stages of activation and localized to the cytosolic or membrane fractions. Antigens with apparent molecular weights of 109, 67, and 48 kd are expressed after priming with IFN-gamma whereas LPS induces the enhanced expression of antigens with molecular weights of 46, 30, and 14.4 kd. One antigen with a molecular weight of 54 kd apparently is only expressed by fully activated BMM phi treated with a combination of IFN-gamma and LPS. Antigens with molecular weights of 170 and 21 kd are repressed by LPS and IFN-gamma respectively. IFN-gamma partially counteracts changes induced by treatment with LPS alone when used in combination with LPS. All antigens are localized in the cytosolic fraction except the 54 kd and to some extent the 30 kd, which were detected in the membrane fraction. The 21 kd was only detectable in crude lysates and thus presumably is of nuclear origin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Sharing of the IL-2 receptor {gamma} chain with the functional IL-9 receptor complex

The third subunit, the so-called common (_c) chain, of the IL-2receptor is shared among the receptors for IL-2, IL-4, IL-7and IL-15, and dysfunction of the _c chain is thought to causeX-linked severe combined immunodeficiency (XSCID) ascribed toimpairment of early T cell development. However, cytokines linkedto XSCID are as yet unidentified. A mAb specific for the _c chain,TUGm2, profoundly inhibited cell proliferation in response toIL-9. Another mAb, TUGm3, immunoprecipltated [¹²⁵I]IL-9 cross-linkedwith either the IL-9 receptor or the _c chain. These resultsdemonstrate that the _c chain is included in the functional receptorcomplex for IL-9, which was initially characterized as a T cellgrowth factor and is essential for IL-9-dependent growth signaltransductlon.     

Mechanisms regulating the expression of indoleamine 2,3-dioxygenase during decidualization of human endometrium

BACKGROUND: Expression of the tryptophan catabolizing enzyme, indoleamine 2,3-dioxygenase, in the mouse placenta has been shown to be critical in preventing immunological rejection of the fetal allograft. To clarify the physiological importance of indoleamine 2,3-dioxygenase in human pregnancy, we have studied how the expression of this enzyme changes during decidualization of human endometrium at both the cell and tissue level. METHODS AND RESULTS: The level of indoleamine 2,3-dioxygenase mRNA expression (determined by RT-PCR) was higher in decidual than in endometrial tissue. Uterine decidual tissue in ectopic pregnancy similarly showed increased mRNA expression. Immunohistochemistry demonstrated that indoleamine 2,3-dioxygenase protein immunoreactivity was found in glandular epithelium and in stromal cells. The intensity of this immunoreactivity was increased in decidualized tissue. In a cell culture model, the level of indoleamine 2,3-dioxygenase mRNA was suppressed specifically by progesterone-induced decidualization of isolated endometrial stromal cells. Indoleamine 2,3-dioxygenase protein abundance (determined by Western blot) was also decreased by progesterone-induced decidualization. However interferon-gamma, a potent stimulator of indoleamine 2,3-dioxygenase gene expression, increased the level of indoleamine 2,3-dioxygenase mRNA and protein in both non-decidualized and in decidualized cells. Indoleamine 2,3-dioxygenase activity (determined by measuring the concentration of tryptophan and its indoleamine 2,3-dioxygenase catabolite, kynurenine) was also decreased by progesterone-induced decidualization but enhanced following interferon-gamma treatment. Expression of other interferon-gamma inducible genes (STAT1 and tryptophanyl-tRNA synthetase) showed the same pattern as that of indoleamine 2,3-dioxygenase in tissue samples, but was not changed by decidualization in the cell culture model. CONCLUSIONS: These data suggest that despite suppression by progesterone, indoleamine 2,3-dioxygenase expression in endometrial stromal cells may increase during decidualization due to stimulation by interferon-gamma secreted by infiltrating leukocytes.     

Comprehensive phenotypic analysis of the gut intra-epithelial lymphocyte compartment: perturbations induced by acute reovirus 1/L infection of the gastrointestinal tract

Intestinal intra-epithelial lymphocytes (IELs) form a highly specialized lymphoid compartment. IELs consist primarily of T cells that are dispersed as single cells within the epithelial cell layer that surrounds the intestinal lumen. These lymphocytes along with lamina propria lymphocytes are considered to play an important role in the regulation of immune responses. IELs are heterogeneous with regard to phenotype, and they contain sub-populations with diverse functions. In our most recent study, we found that intra-duodenal inoculation of mice with reovirus serotype 1/strain Lang (reovirus 1/L) induced expression of both germinal center and T cell antigen and CD11c on IELs suggesting these cells to be the recently stimulated cells in gut mucosal tissue. We also demonstrated that IELs from these mice when cultured in vitro in the presence of reovirus 1/L-pulsed antigen-presenting cells generated reovirus 1/L-specific MHC-restricted CTL whose function was mediated utilizing perforin, Fas-FasL and TRAIL mechanisms. This present study provides a comprehensive analysis of the diverse subsets of IELs, which function with other mucosal cells to provide a strong, protective immunity in a highly regulated fashion inside the microenvironment of the intestinal epithelium. We demonstrated that the IEL population contains both thymus-dependent (TD) and thymus-independent (TI) lymphocytes in mice and that a complex phenotype is present when sub-populations are analyzed for TCR, Thy-1, CD4, CD8 and B220 expression in a comprehensive manner. In reovirus 1/L-inoculated mice, we found a decrease in the TI population and an increase in the TD population characterized by significant alterations in various sub-populations. This increase was largely due to an increase in CD4(+), CD8(+) and CD4/CD8 double-positive sub-populations of TD IELs. Intracellular cytokine analysis demonstrated induction of IFN-gamma and an increase in effector/cytotoxic CD8 and CD4 cells after reovirus 1/L infection. These results suggest that TD IELs may play an important role in the clearance of reovirus 1/L infection from gut.     

Distribution of fetal erythroblasts enriched from maternal blood in multifetal pregnancies

BACKGROUND: The aim of this study was to establish the frequency of fetal cells in the maternal blood of multifetal pregnancies and compare this figure with singleton pregnancies. METHODS: We obtained maternal blood from 31 pregnancies with 2-6 fetuses at 11-16 weeks gestation and from 50 normal singleton controls (11-14 weeks gestation). Fetal erythroblasts were isolated from maternal blood using triple density gradient separation and anti-CD71 magnetic cell-sorting techniques. The enriched erythroblasts were stained with Kleihauer-Giemsa and with fluorescent antibodies for the zeta (zeta), epsilon (epsilon) and gamma (gamma) globin chains. The percentage of fetal cells positive for each stain was calculated. Fluorescence in-situ hybridization (FISH) for X and Y chromosomes was also performed. RESULTS: The percentage of erythroblasts enriched from maternal blood that stained positive for zeta, epsilon and gamma globin chains and with Kleihauer-Giemsa was significantly higher in the multifetal compared with singleton pregnancies. The median enriched percentage of positively stained erythroblasts was about three times higher in the twin than in singleton pregnancies (P < 0.0001), nearly twice as high in the triplet than in twin pregnancies (P < 0.01) and five times higher in the triplet than singleton pregnancies (P < 0.0001). FISH for Y chromosome confirmed the increase in fetal cell proportion in the multifetal pregnancies. CONCLUSIONS: These findings suggest that there is an increase in the physiological feto-maternal cell trafficking in multifetal pregnancies compared with singleton pregnancies, which is likely to be due to the increased placental surface area and vasculature.     

In vitro and in vivo recovery of IFN-{gamma}, but not IL-2, production by IL-12 in mice with plasma ceIL tumors

We have previously found that T ceILs from mice bearing plasmaceIL tumors (PCT mice) demonstrate decreased proliferation asweIL as decreased production of the T^h 1-associated cytokinesIL-2 and IFN- in response to polyclonal stimulation. In thepresent study, we have examined soluble factors as possibleelements required to rescue this decreased proliferation andcytokine production by splenocytes from PCT mice. We find thatthe addition of supernatants from stimulated normal splenocyteshas no effect on proliferation or IL-2 production by splenocytesfrom PCT mice. In contrast, these supernatants completely restoreIFN- production by splenocytes from PCT mice. We have foundthat IL-12 is responsible for the observed increase in IFN-production because: (i) addition of anti-IL-12 antibody blocksthis recovery of IFN- production by these supernatants, (ii)the addition of recombinant IL-12 to cultures of splenocytesfrom PCT mice results in increased IFN- production and (iii)In vivo treatment of PCT mice in IL-12 also results in increasedIFN- production by the subsequently activated splenocytes, buthas little effect on proliferation or IL-2 production. Theseresults demonstrate that both in vitro and in vivo, IL-12 selectivelyrestores the decreased production of IFN- by splenocytes fromPCT mice.     

Immunology: Autoimmunity to spermatozoa, asymptomatic Chlamydia trachomatis genital tract infection and {gamma}{delta} T lymphocytes in seminal fluid from the male partners of couples with unexplained infertility

The relationship between an undetected, asymptomatic Chlamydiatrachomatis genital tract infection, the concentration of andb T cells in semen and sperm autoimmunity was examined in 48male partners of couples with unexplained infertility. ImmunoglobulinA (IgA) antibodies to C.trachomatis were detected in seminalfluids from 14 (29.2%) of the men. Only four of these were positivefor circulating anti-chlamydial IgA, suggesting that the stimulusfor antibody production was within the genital tract. In contrast,four men were positive for anti-chlamydial IgG in their semen;all were also seropositive for anti-chlamydial IgG. T lymphocytesbearing the and antigen receptors were present in every semensample. Men with seminal anti-chlamydial IgA, however, had significantly(P = 0.035) elevated semen T cell concentrations (median 3100cells/ml) than did men lacking this antibody (median 1400 cells/ml);concentrations of T cells were comparable in both groups. Genitaltract sperm autoimmunity, as shown by antibodies bound to motileejaculated spermatozoa, was detected in 13 (27.1%) men. Thepresence of these antibodies was associated with elevated concentrationsof both (median 4200 versus 700 cells/ml) and (median 5000versus 850 cells/ml) T cells (P = 0.0002 and 0.0001 respectively).Men with antisperm antibodies only in their serum had seminalT cell concentrations comparable with men testing negative forantisperm antibodies. Anti-chlamydial IgA was identified insemen from four of 10 men with IgA bound to their spermatozoaand in none of the men with only spermatozoabound IgG. Therewas no relationship between sperm quality and the occurrenceof seminal IgA antibodies to either C.trachomatis or spermatozoa.An asymptomatic C.trachomatis infection activates T cells withinthe male genital tract, which may lead to antisperm antibodyformation and immune-mediated infertility.     

Polymorphism in the peroxisome proliferator-activated receptor-gamma gene in women with polycystic ovary syndrome

BACKGROUND: In view of the strong evidence implicating peroxisome proliferator-activated receptor-gamma (PPARgamma) in adiposity and insulin resistance a study was carried out to investigate PPARgamma genotype frequencies in women with polycystic ovary syndrome (PCOS) and to elucidate its role in the pathogenesis of the syndrome. METHODS: The study involved 135 women with PCOS and 115 healthy control women who were genotyped for a known functional variant of the PPARgamma gene using single strand conformation polymorphism (SSCP) analysis. RESULTS: A significantly different allele distribution of the Pro12 Ala polymorphism of the PPARgamma gene was observed between the two groups, with the frequency of the variant Ala isoform being significantly reduced in the PCOS group (12.6%) when compared with the control group (19.1%) (P = 0.045), at an odds ratio of 0.609 (95% confidence interval: 0.374-0.991). The genotype distributions of the Pro12 Ala polymorphism in the PCOS and control groups were different with borderline significance (P = 0.051). CONCLUSIONS: Our data support a role for PPARgamma gene polymorphism in the pathogenesis of PCOS, the presence of the Ala isoform being protective against the development of PCOS.     

A role for BP-3/BST-1 antigen in early T cell development

In the mouse thymus, pre-T cells are defined by their CD3^–CD4^–CD8^–triple-negative, CD44^10/– CD25⁺ phenotype. We made a ratmAb IF-7, that, among all Tcell subsets analyzed, reacted exclusivelywith pre-T cells. Molecular cloning revealed that the antigenrecognized by IF-7 was identical to BP-3/BST-1, a glycosyl-phosphatidylinositol-linked,CD38-related molecule previously described asa possible co-activationmolecule of pre-B cells. We found that IF-7 cross-linking enhancesthe proliferative response ofsorted pre-T cells to anti-CD3stimulation. In addition, IF-7 enhances and accelerates thedevelopment of fetal thymic organ culture (FTOC), although the lineage is unaffected by the treatment. In addition, sortedIF-7⁺ pre-T cells give preferentially rise to ß TCR⁺thymocytes in FTOC. Our observations strongly suggest that BP-3/BST-1is implicated in both early B and T cell growth and development,and is an early marker for the ß lineage.     

A large proportion of bovine T cells express the gamma delta T cell receptor and show a distinct tissue distribution and surface phenotype

The numbers, phenotype, and tissue distribution of gamma delta T cells in cattle were studied using two monoclonal antibodies (mAbs) which react with the bovine gamma delta T cell receptor (TCR). Both mAbs stained 20-40% of T cells in peripheral blood, and immunoprecipitated molecules of 44 and 36 kd (reduced) and 70-80 kd (non-reduced). In cattle the majority of circulating gamma delta T cells showed a distinct surface phenotype; they expressed T19, a 215 kd molecule described in sheep and cattle which marks only gamma delta T cells. Bovine gamma delta T cells were also CD2-, CD4-, and mostly CD8-, and failed to express CD6, a molecule possibly involved in T cell activation. The distribution of gamma delta T cells in cattle lymphoid tissues differed markedly from that in humans, in that bovine gamma delta T cells were concentrated around lymph node trabeculae and were usually sparse or absent from the B cell and T cell domains of lymph nodes. Like most other species studied, gamma delta T cells in cattle were localized to epithelial surfaces, particularly within the skin and intestine, indicating that it was at these sites where gamma delta T cells functioned. Our results provide further evidence for the unusual localization, recirculation pattern, and phenotype of gamma delta T cells, and also show that some features of gamma delta T cells can differ quite markedly from species to species.     

The appearance and role of gamma delta T cells in the peritoneal cavity and liver during primary infection with Listeria monocytogenes in rats.

We have previously reported that gamma delta T cells play important roles in protection during the early stage of infection with Listeria monocytogenes in mice. To generalize the protective roles of gamma delta T cells in listerial infection to different species, we examined the appearance of gamma delta T cells during infection with L. monocytogenes in Fisher F344 rats. The numbers of bacteria in the peritoneal cavity and liver increased to a maximum level on day 3 and then decreased to an undetectable level by day 10 after an intraperitoneal infection with a sublethal dose (1 x 10(8)) of viable L. monocytogenes in rats. CD3+ alpha beta- T cells in the peritoneal cavity and liver began to increase on day 3, reached a maximum level on day 6, and thereafter decreased gradually by day 10 after infection. Northern blot analysis confirmed that the CD3+ alpha beta- T cells expressed TCR delta and gamma gene messages. In vivo treatment with anti-TCR alpha beta mAb, which suppressed most of the alpha beta T cells in the periphery and impaired resistance during the late stage of listerial infection, did not affect the host defense by day 6 after infection. A significantly increased number of gamma delta T cells was detected in the peritoneal cavity of the TCR alpha beta-suppressed rats on day 6 after infection. These results suggest that the early appearing gamma delta T cells may contribute to the host defense at a relatively early stage during listeriosis in rats.     

Interleukin 1alpha and tissue-lytic matrix metalloproteinase-1 are elevated in ectopic endometrium of patients with endometriosis

BACKGROUND: Matrix metalloproteinases (MMP) play an essential role in tissue remodelling and menstruation and appear to be regulated by cytokines such as interleukin-1alpha (IL-1alpha). In order to investigate their role in the pathogenesis of endometriosis, the aim of the present study was to compare the protein localization of matrix metalloproteinase-1 (MMP-1) and of its main stimulatory cytokine IL-1alpha in eutopic and dystopic endometrium of patients with endometriosis. METHODS: MMP-1 and IL-1alpha protein localization was analysed retrospectively in paired paraffin-embedded tissue biopsies obtained simultaneously from the endometrial cavity and from endometrial lesions of 37 patients with peritoneal or ovarian endometriosis and in cycling endometria from 37 women without endometriosis. Protein localization was demonstrated by immunohistochemistry; antibody specificity was confirmed by western blot analysis. RESULTS: MMP-1 and IL-1alpha protein staining in women suffering from endometriosis was significantly more pronounced in endometriotic lesions than in eutopic endometrium. This held true for both epithelial MMP-1 and IL-1alpha staining (P < 0.006 and P < 0.001), and for stromal MMP-1 and IL-1alpha staining (P < 0.001 and P < 0.001). Furthermore, stromal MMP-1 and IL-1alpha were significantly co-expressed in dystopic endometriotic tissue (P = 0.045). Endometrial MMP-1 and IL-1alpha protein expression pattern in eutopic endometrium from women suffering from endometriosis, however, did not differ significantly from the pattern seen in healthy women. CONCLUSIONS: The increased expression of both matrix-degrading MMP-1 and its major stimulatory cytokine IL-1alpha in endometriotic lesions and the selective co-expression in the stroma of endometriotic foci clearly suggests their involvement in the pathogenic mechanisms leading to local invasion and tissue destruction.     

Induction of human hsp60 expression in monocytic cell lines.

Both bacterial and mammalian heat shock proteins (HSP) are recognized by some T cells, and hsp60 recognition has been implicated in rheumatoid arthritis. We have developed a model to study the induction of hsp60 in human monocytic cell lines. An anti-mycobacterial hsp65 mAb (ML30), cross-reacting with human hsp60 was used to screen 21 human tumor cell lines in Western blot analysis. All T cell and B cell lymphomas constitutively expressed hsp60 protein at moderate to high levels, while little or no hsp60 protein was detected in two monocytic leukemia lines. Moderate to high levels of hsp60 mRNA and protein could be induced in the THP-I monocytic leukemia cell line by heat shock, retinoic acid, interferon (IFN)-gamma or tumor necrosis factor (TNF)-alpha treatment, the highest levels obtained with a combination of IFN-gamma/TNF-alpha. This was also seen using two rabbit anti-hsp60 antisera directed against the N-terminal or C-terminal part of the human hsp60 protein. The determinants detected by the ML30 mAb or the two rabbit anti-hsp60 antisera were not cell surface expressed, as measured with immunofluorescence (FACS) analysis on control cultured or cytokine treated cell lines. This could be a useful model for studies related to the induction of hsp60 in human cells.