Vectorial production of interleukin 1 and interleukin 6 by rat Sertoli cells cultured in a dual culture compartment system

The bidirectional production of interleukin-1 (IL-1) and IL-6 by Sertoli cells and its regulation by inflammatory and physiological stimuli has been studied using a dual compartment culture system allowing the study of Sertoli cell apical and basal secretory activities. Another Sertoli cell activity, the vectorial transferrin production was also studied in all culture conditions. A low constitutive IL-1 production appeared equally distributed between both poles, while IL-6 and transferrin constitutive production was predominantly directed apically. Two activators of macrophages, lipopolysaccharides and zymosan, were found to induce marked increases of IL-1 in the compartment where they had been added: basal if added to the lower compartment and vice versa. In contrast, after a basal stimulation, IL-6 production was mainly increased in the upper compartment that corresponds to a Sertoli cell apical flux. In this system, IL-1 and IL-6 levels were not modified by FSH; they were not also affected by residual bodies and latex beads, probably due to the fact that, in the bicameral system, phagocytosis is restricted to the Sertoli cells situated at the surface of the inner compartment. IL-1beta, but not IL-1alpha, induced IL-6 secretion in the compartment of stimulation. In conclusion, the present study demonstrates that vectorial secretory patterns of IL-1 and IL-6 production greatly differ and that these cytokines are also differently regulated. These results suggest that Sertoli IL-1 and IL-6 have different targets within the testis and that, in normal and pathophysiological conditions, both the tubular and the interstitial compartments may be influenced by the action of these paracrine factors.     

Erythromycin inhibits tumor necrosis factor alpha and interleukin 6 production induced by heat-killed Streptococcus pneumoniae in whole blood

To determine the effects of penicillin and erythromycin on cytokine production induced by heat-killed Streptococcus pneumoniae (HKSP), we studied the effects of those drugs on cytokine production induced by S. pneumoniae in human whole blood in vitro and ex vivo. In whole blood in vitro, erythromycin, but not penicillin, caused a dose-dependent decrease in HKSP-induced production of tumor necrosis factor alpha (TNF) and interleukin 6 (IL-6), while the production of IL-10, IL-12, and gamma interferon was inhibited only at the highest erythromycin concentration tested (10(-3) M). The production of TNF and IL-6 in whole blood obtained from healthy subjects after a 30-min infusion of erythromycin (1,000 mg) was lower after ex vivo stimulation with HKSP than that in blood drawn before the infusion. Inhibition of TNF contributed to erythromycin-induced inhibition of IL-6 synthesis. Inhibition of TNF and IL-6 production by erythromycin may have a negative impact on host defense mechanisms during pneumococcal pneumonia.     

Apoptosis induction and potent antiestrogen receptor-negative breast cancer activity in vivo by a retinoid antagonist

Close to 180,000 women will be diagnosed with breast cancer this year in the United States and more than 43,000 will die from this disease. Antiestrogens have shown promise, but they can only be effective against estrogen-dependent stages of the disease. We identify here a retinoid antagonist, MX781, that is effective against estrogen receptor-positive and -negative breast cancer cells. Although classical retinoids show limited efficacy and significant side effects, this novel compound kills breast cancer cells by inducing apoptosis and is effective against estrogen receptor-negative human breast cancer tumors in vivo. Remarkably, MX781 is well tolerated and does not seem to have significant toxicity. This novel retinoid antagonist, therefore, represents a promising new candidate for the treatment of breast cancer.     

Stimulation by prostaglandin F2alpha of acidic glycosaminoglycan production in cultured fibroblasts

The effect of PGF2alpha on the synthesis of hexosamine-containing substances (acidic glycosaminoglycans and glycoproteins) was studied in cultured fibroblasts derived from a rat carrageenin granuloma. Treatment with PGF2alpha ranging from 0.01 mug/ml to 20 mug/ml resulted in a significant increase of the production of these macromolecules by the cells. The stimulatory effect was found significant even at the low concentration of 10 ng/ml, and could be seen as early as 3h after exposure to PGF2alpha. The hexosamine-containing substances increased by PGF2alpha revealed that 80% of the increase was due to acidic glycosaminoglycans and the rest was due to glycoproteins.     

Induction of mitogen-inducible nuclear orphan receptor by interleukin 1 in human synovial and gingival fibroblasts

High levels of interleukin 1 (IL-1) found in inflammatory diseases such as rheumatoid arthritis and periodontitis act on the local fibroblasts, resulting in an altered phenotype characterized by hyperplasia and the production of inflammatory mediators and destructive enzymes. The goal of this study was to identify genes induced as an early response to IL-1 in synovial and gingival fibroblasts which might play a regulatory role in the cascade of events leading to their activation. Using the technique of mRNA differential display, we have identified the mitogen-inducible nuclear orphan receptor (MINOR) as a gene up-regulated by IL-1 in human synovial and gingival fibroblasts. The rapid induction of both mRNA and DNA binding activity suggests that MINOR may play an important early role in regulating the response of fibroblasts to inflammation.     

Effects and interactions of tumour necrosis factor alpha and bradykinin on interleukin-1 production in gingival fibroblasts

We examined the association of serum albumin concentration with diabetes mellitus and other cardiovascular risk factors, prevalent cardiovascular disease, and ultrasonographically assessed carotid artery intima-media thickness using data from 45- to 64-year-old adults in the Atherosclerosis Risk in Communities (ARIC) Study. The mean albumin concentration was 0.04 to 0.12 g/L lower in participants with diabetes and 0.02 to 0.06 g/L lower in those with cardiovascular disease, compared to participants without these conditions. However, lower serum albumin level was also correlated with most traditional risk factors and hemostatic variables. On adjustment for these, there was essentially no association between serum albumin and prevalent cardiovascular disease. Likewise, there was no association between albumin and carotid intima-media thickness (a marker of atherosclerosis). While hypoalbuminemia may be a marker for chronic disease and perhaps renal loss of albumin, it seems unlikely that it is an important cause of atherosclerosis.     

Effects of human immunodeficiency virus and colony-stimulating factors on the production of interleukin 6 and tumor necrosis factor alpha by monocyte/macrophages

Patients infected with human immunodeficiency virus (HIV) frequently have increased production of interleukin 6 (IL-6) and tumor necrosis factor alpha (TNF-alpha), and these cytokines may in turn contribute to the disease pathogenesis. It has been hypothesized that secretion of these cytokines by HIV-exposed mononuclear cells or HIV-infected monocyte/macrophages (M/Ms) is the principal source of their overproduction in HIV-infected patients, and the present study was undertaken to explore this issue. We observed that in the absence of endotoxin or cytokines, M/Ms productively infected by HIV do not produce detectable IL-6 or TNF-alpha. However, granulocyte-macrophage colony-stimulating factor (GM-CSF), a cytokine that enhances HIV replication in M/Ms and is frequently used to propagate monocytotropic strains of HIV, can induce the relatively long-term production of IL-6 (up to 47 U/ml) and TNF-alpha (up to 47 pg/ml) by M/Ms, even in the absence of HIV. Also, HIV induced production of a relatively small (< or = 9 U/ml) quantity of IL-6 in M/Ms stimulated with macrophage-colony stimulating factor (M-CSF). Finally, while highly concentrated HIV induced production of both cytokines by either M/Ms or peripheral blood mononuclear cells (PBMCs), this production was almost completely eliminated when care was taken to avoid contamination of HIV by endotoxin. These data suggest that the excess IL-6 and TNF-alpha in HIV-infected patients does not simply result from their production by HIV-infected M/Ms and that alternative mechanisms are involved in this process.     

A randomized phase II trial of interleukin 2 and interleukin 2-interferon alpha in advanced renal cancer

An 81 year old right handed woman developed a left alien hand syndrome characterised by involuntary movements of choking and hitting the face, neck, and shoulder. The patient showed multiple disorders of primary sensation, sensory processing, hemispatial attention, and visual association, as well as a combination of sensory, optic, and cerebellar ataxia (triple ataxia) of the left arm in the absence of motor neglect or hemiparesis. Imaging studies disclosed subacute infarction in the right thalamus, hippocampus, inferior temporal lobes, splenium of corpus callosum, and occipital lobe due to right posterior cerebral artery occlusion. This rare syndrome should be considered as a "sensory" or "posterior" form of the alien hand syndrome, to be distinguished from the "motor" or "anterior" form described more commonly.     

Autologous versus allogeneic T cell-stimulated IL-6 production by dermal fibroblasts

T cells adhere to human dermal fibroblasts (HDF). This cellular interaction leads to a pronounced secretion of the proinflammatory cytokines IL-6 and IL-8 via a juxtacrine stimulation induced by HDF-associated IL-1. Upon stimulation, fibroblasts express various surface proteins such as MCH-I molecules, which may interact with corresponding receptors on T cells. The present study was conducted to further investigate the mechanism of this complex interaction with regard to the secretion of IL-6 in cocultures of T cells and HDF. IL-6 was time- and dose-dependently upregulated in such cocultures. Spatial separation of the cells by microporous membranes resulted in a 90% reduction of IL-6 secretion, but when cells had limited cell contact IL-6 secretion was increased again. Allogeneic cocultures of T cells and HDF showed increased capacity of IL-6 stimulation as compared to autologous cultures. Our results suggest that MHC-I/T cell receptor interaction modulates IL-6 secretion in allogeneic and autologous cocultures.     

Interleukin-1alpha and tumor necrosis factor alpha synergistically stimulate prostaglandin E2-dependent production of interleukin-11 in rheumatoid synovial fibroblasts

OBJECTIVE: Interleukin-11 (IL-11), an IL-6-type cytokine, is thought to be involved in bone resorption via osteoclast differentiation. Here, we characterized the combined effect of IL-1alpha and tumor necrosis factor alpha (TNFalpha), major cytokines in the rheumatoid synovium, on the production of IL-11 by cultured rheumatoid synovial fibroblasts (RSFs). METHODS: The amounts of IL-11, IL-6, and prostaglandin E2 (PGE2) were measured by enzyme-linked immunosorbent assay. IL-11 messenger RNA (mRNA) levels were determined by Northern blotting. Protein expression of cytosolic phospholipase A2 (cPLA2), cyclooxygenase 2 (COX-2), and protein kinase C (PKC) isoforms were determined by Western blotting. RESULTS: IL-1alpha and TNFalpha synergistically stimulated RSFs to produce IL-11 at both the mRNA and protein levels. This synergistic effect was completely inhibited by indomethacin. The inhibition was prevented by PGE2, indicating that the synergistic effect of IL-1alpha and TNFalpha was PGE2-mediated. The cooperative effects of these 2 cytokines were also observed in the production of PGE2 and the expression of 2 regulatory enzymes in PGE2 production, cPLA2 and COX-2. The synergistic induction of IL-11 by IL-1alpha and TNFalpha was completely inhibited by a potent inhibitor of all isoforms of PKC, GF109203X. In contrast, phorbol myristate acetate, which induced a down-regulation of PKC, degrading all PKC isoforms except atypical PKC, did not affect the induction of IL-11. CONCLUSION: These findings suggest that IL-1alpha and TNFalpha synergistically stimulate the production of IL-11 via their effects on PGE2 production in the rheumatoid joint, and that atypical PKC may be another target for down-regulation of IL-11, the bone resorption-associated cytokine.     

Advanced colorectal cancer is associated with impaired interleukin 12 and enhanced interleukin 10 production

Interleukin 12 (IL-12) is a heterodimeric cytokine that has been demonstrated to have a major role in stimulating a cell-mediated antitumor response. IL-10, a product of T helper 2 lymphocytes, is its most potent inhibitor. The aim of this study was to investigate whether patients with colorectal cancer had an imbalance in production of IL-12 and IL-10 preoperatively, and whether this was associated with advanced disease at surgery. Blood was obtained before surgery from 60 patients with colorectal cancer and from 30 controls. Peripheral blood mononuclear cells were incubated with Staphylococcus aureus Cowan's strain 1 in vitro for 24 h to assess IL-12 expression after stimulation, and serum was used for IL-10 measurement. IL-12 and IL-10 levels were assessed by ELISA. A single pathologist staged the tumors according to the tumor-node-metastasis (TNM) and Dukes' classifications. Patients with colorectal cancer had significantly lower levels of IL-12 (P <0.001) and higher levels of IL-10(P = 0.004) compared to controls. In addition, lower levels of IL-12 were detected in those patients who were node positive (P<0.05), had Dukes' C lesions (P < or = 0.001), and T3 or T4 lesions (P<0.033) when compared to controls. Patients with Dukes' B and C lesions (P<0.01) and T3 and T4 lesions (P<0.05) also had higher levels of IL-10 compared to controls. This study is the first to demonstrate that patients with colorectal cancer have decreased IL-12 production and increased serum IL-10. This suggests an impaired T helper 1 cell-mediated antitumor response and provides some justification for exogenous IL-12 therapy or anti-IL-10 therapy in these patients.     

Models of estrogen receptor regulation by estrogens and antiestrogens in breast cancer cell lines

The expression and stability of the estrogen receptor (ER) is the result of a complex process that is modulated by estrogens and antiestrogens. Regulation of the steady-state ER mRNA and protein levels in breast cancer cells appears to be the result of either of two distinct regulatory mechanisms. Estrogen exposure causes a rapid down-regulation of the steady-state level of ER mRNA and protein in model I regulation, as exemplified by the MCF-7:WS8 cell line. Conversely, in model II regulation, as observed in the T47D:A18 cell line, estrogen exposure causes an increase in the steady-state ER mRNA level and a maintenance of the ER protein level. In both these cell lines, the nonsteroidal antiestrogen 4-hydroxytamoxifen has little effect on the mRNA level but causes a net accumulation of the ER protein over time. In contrast, the pure antiestrogen ICI 182,780 causes a dramatic reduction of the ER protein in both the MCF-7:WS8 and T47D:A18 cell lines. This loss has little effect upon the ER mRNA level in the MCF-7:WS8 cells but leads to a decline in the ER mRNA in the T47D:Al8 cells. The estrogen-independent MCF-7:2A cell line, which has adapted to growth in estrogen free media, expresses two forms of the ER, a wild-type Mr66,000 ER and a mutant Mr77,000 ER (ER77). ER77 is the product of a genomic rearrangement resulting in a tandem duplication of exons 6 and 7 (J. J. Pink et al, Nucleic Acids Res., 24:962-969,1996). This exon duplication has abolished ligand binding by this protein. Here we demonstrate that the loss of ligand binding has eliminated the effects of 4-OHT and ICI 182,780 on the steady-state ER77 protein level. However, in the MCF-7:2A cells, antiestrogens affect the wild-type ER protein in the same manner as observed in the MCF-7:WS8 and T47D:A18 cells. Estrogen regulates the ER mRNA and wild-type ER and ER77 proteins in the MCF-7:2A cells in the same manner as observed in the MCF-7:WS8 cells. Interestingly, treatment of the MCF-7:2A cells with ICI 182,780 causes a slight increase in ER mRNA, which is reflected in a net increase in the ER77 protein but a dramatic decrease in the wild-type ER. The models presented here describe the response of two human breast cancer cell lines in short-term studies. These distinct regulation pathways are predictive of the response of these cell lines to long-term estrogen deprivation. This study illustrates two alternative regulation pathways that are present in ER-positive, estrogen-dependent breast cancer cells. This variable response highlights the diversity of responses potentially present in the heterogeneous cell populations of clinically observed breast cancer.