Failure to induce nitric oxide production by human monocyte-derived macrophages. Manipulation of biochemical pathways

Production of nitrix oxide (NO-) by human macrophages is controversial. In the present study, the ability of human monocyte-derived macrophages (M phi) to produce NO- in response to M phi modulators was tested. M phi cultured for up to nine days and stimulated for 48 with different concentrations of LPS and/or IFN-gamma failed to produce significant amounts of NO2- compared to unstimulated cultures. Inhibition of the cyclo-oxygenase pathway with indomethacin did not increase NO2- production by LPS stimulated M phi. Since human M phi lack biopterin, needed for NO- synthesis by murine M phi, human M phi stimulated with LPS plus IFN-gamma were additionally cultured in the presence of neopterin or biopterin. These treatments did not induce NO2- production. Furthermore, simultaneous treatment with indomethacin and neopterin or biopterin also failed to induce NO2- production. However, human M phi, stimulated with IFN-gamma and LPs, produced TNF-alpha suggesting that the lack of increment in NO2- production was not due to an absence of response of M phi to the stimuli used. As an indirect approach to explore the NO- production, human M phi were infected with virulent Mycobacterium tuberculosis H37Rv and simultaneously treated with the competitive inhibitor NGmonomethyl-L-arginine (NGMMA). Mycobacterial intracellular replication was measured by 3H-uracil incorporation. NGMMA did not have any effect on mycobacterial replication. These results further suggest that human M phi do not produce NO- at least by the inducible pathway.     

Role of macrophages in regeneration of liver

In an attempt to clarify the role of macrophages and their mediators during regeneration of the liver, the difference of liver regeneration among C3H/HeN (LPS-responsive strain) and C3H/HeJ (LPS-resistant strain) mice was investigated. After a 67% partial hepatectomy, an increase in the weight of regenerating liver was significantly delayed in the C3H/HeJ mice, as compared with C3H/HeN mice. The number of hepatocytes labeled with antibody against PCNA reached maximum levels 48 hr after partial hepatectomy, but the PCNA labeling index in C3H/HeJ mice was 20% less than that for C3H/HeN mice. In addition, TNF-alpha activities in serum were enhanced shortly after partial hepatectomy in C3H/HeN strain mice, but were not increased in C3H/HeJ strain mice. Serum IL-6 levels were markedly enhanced in both C3H/HeN and C3H/HeJ mice, but a bimodial peak (14 and 48 hr after partial hepatectomy) was demonstrated in C3H/HeN mice, in contrast to a single peak (at 24 hr) in C3H/HeJ mice. Suppression of Kupffer cells by previous administration of gadolinium chloride in C3H/HeN mice reduced the increase in both serum TNF-alpha and IL-6 concentrations, reduced PCNA labeling index of hepatocytes by 20%, and disturbed the regeneration of the liver. Previous administration of antibody against TNF-alpha reduced the PCNA labeling index of hepatocytes by 20% after partial hepatectomy in C3H/HeN strain mice. These results suggest that LPS-responsive macrophages in the liver and their mediators, especially TNF-alpha, could partly play a role in liver regeneration.     

Significant contribution of spleen cells in mediating the lethal effects of endotoxin in vivo

Two closely related, histocompatible mouse strains that have marked differences in both in vitro and in vivo responses to endotoxin were used to evaluate the contribution of lymphoid cells to the lethal effect of endotoxin. C3H/HeJ mice are endotoxin resistant, whereas C3H/HeN mice are endotoxin sensitive. In vitro spleen cell mitogenic responses to endotoxin were similar in untreated mice and in mice that received sublethal irradiation (450 R) followed by reconstitution with autologous spleen cells. Reconstitution with spleen cells from the related strain produced chimeric animals with spleen cell mitogenic activity like that of the donor strain. When chimeric animals were subjected to a lethal challenge of endotoxin, their response was markedly altered by the transferred lymphoid cells. C3H/HeJ animals reconstituted with C3H/HeN cells became more endotoxin sensitive, whereas C3H/HeJ cells became more endotoxin resistant. These results indicate that spleen cells play a significant, detrimental role in endotoxin-induced lethality.     

Tumor necrosis factor (TNF) induction from monocyte/macrophages by Candida species

Candida albicans was studied for its capacity to induce TNF production from mouse peritoneal macrophages (PM phi). TNF activities in the culture supernatants of Candida-stimulated PM phi and human peripheral blood monocytes were assessed by L 929 bioassay and ELISA respectively. C. albicans induced TNF production from PM phi and human peripheral blood monocytes in a dose-dependent manner. Although the capacity was lesser than live form, heat-killed C. albicans was also found to be capable of stimulating PM phi to induce TNF. The filtered supernatant of 24 h cultured live C. albicans had no effects on TNF production from PM phi. Saccharomyces cerevisiae-extracted mannan, a yeast cell wall antigen, induced TNF production from PM phi in a dose-dependent manner. Thus, the effect of C. albicans on TNF production from PM phi was seemed to be directly related to the presence of the yeast cell wall itself. Compatible with these data, when various candida species (C. albicans, C. tropicalis, C. pseudotropicalis. C. lusitaniae, C. krusei, C. parapsilosis, C. guilliermondii, C. stellatoidea, C. glabrata) and S. cerevisiae were compared to each other, at a concentration of 2 x 10(6) yeast cells/ml from each species, it was observed that TNF inducing capacities varied. Among the species used in this study, C. guilliermondii and C. glabrata, of which the yeast cell size were the smallest ones, were found to be less potent than that of others to induce TNF from PM phi.     

Involvement of CD14 and complement receptors CR3 and CR4 in nuclear factor-kappaB activation and TNF production induced by lipopolysaccharide and group B streptococcal cell walls

This study was undertaken to evaluate the role of CD14 and complement receptors type 3 (CR3) and 4 (CR4) in mediating TNF release and NF-kappaB activation induced by LPS and cell wall preparations from group B streptococci type III (GBS). LPS and GBS caused TNF secretion from human monocytes in a CD14-dependent manner, and soluble CD14, LPS binding protein, or their combination potentiated both LPS- and GBS-induced activities. Blocking of either CD14 or CD18, the common beta-subunit of CR3 and CR4, decreased GBS-induced TNF release, while LPS-mediated TNF production was inhibited by anti-CD14 mAb only. Chinese hamster ovary cell transfectants (CHO) that express human CD14 (CHO/CD14) responded to both LPS and GBS with NF-kappaB translocation, which was inhibited by anti-CD14 mAb and enhanced by LPS binding protein. While LPS showed fast kinetics of NF-kappaB activation in CHO/CD14 cells, a slower NF-kappaB response was induced by GBS. LPS also activated NF-kappaB in CHO cells transfected with either human CR3 or CR4 cDNA, although responses were delayed and weaker than those of CHO/CD14 cells. In contrast to LPS, GBS failed to induce NF-kappaB in CHO/CR3 or CHO/CR4 cells. Both C3H/OuJ (Lps[n]) and C3H/HeJ (Lps[d]) mouse peritoneal macrophages responded to GBS with TNF production and NF-kappaB translocation, whereas LPS was active only in C3H/OuJ macrophages. Thus, LPS and GBS differentially involve CD14 and CR3 or CR4 for signaling NF-kappaB activation in CHO cells and TNF release in human monocytes, and engage a different set of receptors and/or intracellular signaling pathways in mouse macrophages.     

Differences in the shedding of soluble TNF receptors between endotoxin-sensitive and endotoxin-resistant mice in response to lipopolysaccharide or live bacterial challenge

TNF-alpha plays a pivotal role in the pathogenesis of septic shock. It exerts its effects by binding two cell surface receptors, designated TNF-R I and II, also referred to as the p55 and p75 receptors, respectively. TNF-Rs are transmembrane proteins, which on cleavage of their extracellular domains, result in the release of soluble fragments (sTNF-R). sTNF-R levels increase markedly during infection, and may serve to modulate TNF-alpha bioactivity. The mechanisms regulating this process are uncertain. To investigate this, we measured sTNF-R release in endotoxin-sensitive C3H/HeN and endotoxin-resistant C3H/HeJ mice given LPS or live Gram-negative bacteria. In C3H/HeN mice, there was a rapid early response during the first 4 h, and a second peak at 8 h, particularly noticeable in the case of the p75 receptor. Prior administration of neutralizing Abs to TNF-alpha or IFN-gamma had no effect on receptor shedding. Surprisingly, C3H/HeJ mice also responded to both bacterial challenge and to LPS by shedding sTNF-R; the magnitude and duration of the early response was not substantially different from C3H/HeN mice, although the second peak was absent. Peritoneal macrophages from C3H/HeN mice responded promptly (5 h) when stimulated with LPS in vitro, and by 22 h levels had increased five- to 10-fold. In contrast, cells from C3H/HeJ mice demonstrated only a very modest response at 22 h following maximal stimulation. The data suggest that there may be at least two separately regulated pathways that control sTNF-R shedding in these mice.     

Early consequences of macrophage-Francisella tularensis interaction under the influence of different genetic background in mice

The induction, regulation and expression of protective immunity against Francisella tularensis LVS infection is dependent on the results of primary interaction between the cells of host's immunoregulatory system and the microbe. The early events, at least on the side of macrophages, are under the genetic control. To determine the impact of genes that might be involved in the control of resistance to Francisella tularensis LVS infection, we have used three different inbred strains of mice with increasing resistance to this infection in order C3H/HeJ (Lpsd), C3H/HeN (Lpsn"), and C57B1/10N (Lpsn"). The controlled production of IL-10, IFN-gamma, and TNF-alpha coupled with increased production of reactive oxygen metabolites during early phase of infection distinguished less susceptible C3H/HeN mice from their more susceptible cogenic C3H/HeJ counterparts. The enhancement of oxidative metabolism that appeared on day 5 after the infection of both C3H/HeN and C57B1/10N mice closely correlated with increasing resistance of these two strains of mice to Francisella tularensis LVS infection. These mice were also capable to reach the highest level of TNF-alpha on day 5 after the infection. At the same time interval, only C57B1/10N mice produced significantly enhanced level of nitric oxide. Overall, these parameters may suggest their possible biological role in early-phase resistance to Francisella tularensis LVS infection and their subsequent consequences for ultimate control of infection and its clearance.     

Endogenous norepinephrine regulates tumor necrosis factor-alpha production from macrophages in vitro

Evidence for the extraneuronal accumulation of norepinephrine has been demonstrated to occur in macrophage (M phi), yet the physiologic role of this system remains undefined. We have assessed the response of murine peritoneal M phi to adrenergic antagonists. We have also defined a physiologic role of a M phi-associated pool of the nonspecific adrenergic agonist norepinephrine. We investigated the constitutive involvement of alpha-adrenergic and beta-adrenergic receptors in LPS-induced TNF-alpha production. CFA-elicited M phis were incubated with LPS (1 microgram/ml) in the presence or absence of adrenergic agonists and/or antagonists. Although stimulation of alpha-adrenergic receptors increased TNF production and gene expression, beta-adrenergic receptors decreased it. Interestingly, when adrenergic antagonists along with LPS alone were added to M phi, they generated the response opposite to that produced by their suitable agonist, suggesting a role for endogenous norepinephrine in M phi. Thus, although alpha 2-adrenergic antagonists attenuated TNF production, beta-adrenergic antagonists augmented TNF expression in a concentration-dependent manner. Norepinephrine and epinephrine were found in M phi as determined by HPLC and LPS stimulation induced a significant decrease in their content. M phis were also incubated with LPS or medium only, washed, and then challenged 12 h later with LPS. When given a second LPS stimulation, M phis were found to have an increased response to alpha 2-adrenergic agonists and decreased response to alpha 2-adrenergic antagonists. Therefore, M phi-associated norepinephrine appears to regulate LPS-induced TNF production in an autocrine fashion.     

Hemodynamic studies of patients with recent myocardial infarct by means of radiocardiography

The effect of endotoxin to depress the hepatic drug-metabolizing enzyme activity has been studied in the C3H/HeJ and C3H/HeN strains of mice. The C3H/HeJ mouse strain is generally considered to be unresponsive to the biological effects of endotoxin. However, injection of these mice with 0.5 mg/kg body weight of E.coli endotoxin (Westphal extracted) produced a decrease in the rate of N-demethylation of ethylmorphine and in the levels of cytochrome P-450 and cytochrome b5 comparable to that observed in the endotoxin-sensitive C3H/HeJ mouse strain. Although the mechanism of endotoxin action to decrease hepatic microsomal drug-metabolizing activity is presently unknown, the results suggest that: 1) the C3H/HeJ mouse stain is responsive to this endotoxin effect, and 2)that cellular constituents other than B-cells or macrophages are probably involved in eliciting the response, since these cells of the C3H/HeJ mouse are unresponsive to endotoxin.     

In vivo and in vitro evidence for the involvement of tumor necrosis factor-alpha in the induction of leptin by lipopolysaccharide

To examine the role of tumor necrosis factor-alpha (TNF alpha) in mediating leptin secretion during an immunological challenge, we studied the effects of lipopolysaccharide (LPS) and TNF alpha on leptin secretion in endotoxin-sensitive C3H/HeOuJ (OuJ) mice, endotoxin-insensitive C3H/HeJ (HeJ) mice, and primary adipocytes cultured from both. Intraperitoneal injection of LPS increased plasma concentrations of TNF alpha and leptin in OuJ mice, but not in HeJ mice, suggesting a causal relationship between the induction of TNF alpha and leptin. Consistent with this idea, i.p. injection of recombinant murine TNF alpha increased plasma leptin in both OuJ and HeJ mice. To determine whether TNF alpha induces leptin secretion by acting directly on fat cells, primary adipocytes from OuJ and HeJ mice were cultured in the presence of TNF alpha or LPS. Whereas LPS was without effect on leptin secretion by adipocytes, TNF alpha induced a marked increase in the cell supernatant leptin concentration. These data demonstrate that TNF alpha plays a role in regulating the increase in leptin caused by LPS. Moreover, they show that TNF alpha can act directly on adipocytes to stimulate leptin secretion. Our results are consistent with the emerging view that leptin is a key hormone coupling immune system activity to energy balance.     

A novel component different from endotoxin extracted from Prevotella intermedia ATCC 25611 activates lymphoid cells from C3H/HeJ mice and gingival fibroblasts from humans

A novel immunobiologically active fraction was prepared from a phenol-water extract of Prevotella intermedia ATCC 25611 by Sephadex G-100 column chromatography. The fraction consisted mainly of carbohydrate and protein and was devoid of fatty acid. The fraction showed high-molecular-weight bands (10,000 to 12,000) on deoxycholate polyacrylamide gel electrophoresis (DOC-PAGE) and was scarcely active in a Limulus test. We designated the fraction Prevotella glycoprotein (PGP). The PGP fraction showed strong mitogenicity on splenocytes and cytokine-inducing activities on peritoneal macrophages from both C3H/HeJ and C3H/HeN mice, and it stimulated human gingival fibroblasts to produce cytokines. The activities of the PGP fraction were resistant to heat inactivation (100 degrees C for 1 h) and protease treatments and were scarcely inhibited by polymyxin B. In contrast, the purified lipopolysaccharide fraction (LPS-PCP) extracted from the same bacterium with a phenol-chloroform-petroleum ether mixture, which showed strong Limulus activity and a single low-molecular-weight band (approximately 3,000) on DOC-PAGE, lacked the activities on splenocytes and macrophages from C3H/HeJ mice and human gingival fibroblasts. The activities of the LPS-PCP fraction on cells from C3H/HeN mice were completely inhibited by polymyxin B. The LPS extracted from the same bacterium with hot phenol-water (LPS-PW) exhibited the properties of both the PGP fraction and the LPS-PCP fraction. These findings suggest that the unique bioactivities of the LPS-PW fraction of oral black-pigmented bacteria reported to date, which differed from those of the classical endotoxin, were derived from the PGP fraction and not from the LPS itself.     

CD14-dependent internalization of bacterial lipopolysaccharide (LPS) is strongly influenced by LPS aggregation but not by cellular responses to LPS

We analyzed the impact of ligand aggregation and LPS-induced signaling on CD14-dependent LPS internalization kinetics in human monocytic THP-1 cells and murine macrophages. Using two independent methods, we found that the initial rate and extent of LPS internalization increased with LPS aggregate size. In the presence of LPS binding protein (LBP), large LPS aggregates were internalized extremely rapidly (70% of the cell-associated LPS was internalized in 1 min). Smaller LPS aggregates were internalized more slowly than the larger aggregates, and LPS monomers, complexed with soluble CD14 in the absence of LBP, were internalized very slowly after binding to membrane CD14 (5% of the cell-associated LPS was internalized in 1 min). In contrast, the initial aggregation state had little or no effect on the stimulatory potency of the LPS. Previous studies suggest that LPS-induced signal responses may influence the intracellular traffic and processing of LPS. We found that elicited peritoneal macrophages from LPS-responsive (C3H/HeN) and LPS-hyporesponsive (C3H/HeJ) mice internalized LPS with similar kinetics. In addition, pre-exposure of THP-1 cells to LPS had no effect on their ability to internalize subsequently added LPS, and pre-exposure of the cells to the LPS-specific inhibitor, LA-14-PP, inhibited stimulation of the cells without inhibiting LPS internalization. In these cells, LPS is thus internalized by a constitutive cellular mechanism(s) with kinetics that depend importantly upon the physical state in which the LPS is presented to the cell.     

Lipopolysaccharide induction of MARCKS-related protein and cytokine secretion are differentially impaired in microglia from LPS-nonresponsive (C3H/HeJ) mice

Many events involved in activation of microglia and leukocytes by lipopolysaccharide (LPS) are mediated by protein kinase C (PKC), and we have recently demonstrated that a major PKC substrate, MARCKS-related protein (MRP), is selectively induced by LPS in murine microglia. In microglia from LPS-nonresponsive (C3H/HeJ) mice, induction of MRP and secretion of CSF-1 required much higher LPS concentrations (> or = 100 ng/ml) than in normal (C3H/OuJ) microglia (< or = 10 ng/ml). By contrast, TNF alpha production was not significantly increased in C3H/HeJ microglia even at 1 microgram LPS/ml. Microglia expressed PKC isoforms alpha, beta, delta, and zeta (but not gamma and epsilon); PKC isoform levels were similar in both normal and C3H/HeJ microglia and no significant change in response to LPS was noted. Our results indicate that LPS alters PKC substrate (rather than kinase) expression, and that the Lpsd mutation in C3H/HeJ mice differentially affects regulation of several gene products implicated in microglial function.     

The induction of polyclonal B-cell activation and interleukin-1 production by the 75-kDa cell surface protein from Porphyromonas gingivalis in mice

The immunobiological activities of 75-kDa protein, fimbrial protein and lipopolysaccharide lipopolysaccharide prepared from whole cells of Porphyromonas gingivalis 381 were compared. The 75-kDa protein was mitogenic for BALB/c nu/nu, BALB/c and lipopolysaccharide-responsive C3H/HeN mouse spleen cells and for lipopolysaccharide-non-responsive C3H/HeJ mouse spleen cells. The response was significant in BALB/c mouse spleen cells incubated with 1-100 micrograms/ml of the 75-kDa protein. Furthermore, the 75-kDa protein exhibited polyclonal B-cell activation in murine spleen cells, which was similar to the lipopolysaccharide of P. gingivalis. In contrast, fimbriae from P. gingivalis did not, or only weakly, activated murine spleen cells. C3H/HeN mouse macrophages exposed to 10 micrograms/ml of the 75-kDa protein released large amounts of interleukin-1 (IL-1), which were maximal for 48 h, whereas IL-6 activity in macrophage supernatants was not detected throughout the culture period. These results suggest that the 75-kDa protein is a potent polyclonal B-cell activator and that it stimulates IL-1 production from murine peritoneal macrophages as well as lipopolysaccharide, which may play an important part in the inflammatory response during the development of periodontal diseases.     

Histamine synthesis by cells of the macrophage lineage in bone marrow of mice

An injection of Escherichia coli lipopolysaccharide (LPS) increased the activity of histidine decarboxylase (HDC) in bone marrow (BM) cells of C3H/HeN mice much more than in C3H/HeJ mice, which are resistant to various effects of LPS. In WBB6/F1 (W/Wv) mice, which are genetically deficient in mast cells, HDC activity increased more than in C3H/HeN mice. Cultured BM cells of W/Wv mice spontaneously synthesized histamine in a HDC-dependent way. LPS caused a slight increase in HDC-associated histamine synthesis by these cells. Treatment of the BM cells with murine recombinant granulocyte-macrophage colony stimulating factor (mrGM-CSF) increased the histamine synthesis. In addition, treatment with mrGM-CSF made the cells respond to LPS by a dose-dependent increase in HDC activity and histamine synthesis. Most dish-adherent BM cells that had been treated with both mrGM-CSF and LPS for 48 h were stained for nonspecific esterase and not for chloroacetate esterase, and had twice as much HDC activity as the nonadherent cells had. Immunocytochemical analysis of the BM cells of W/Wv mice treated with both mrGM-CSF and LPS showed that HDC was in the cytoplasm of cells having Mac-1, a macrophage-differentiation antigen. These results suggest that cells of the macrophage lineage in the BM of mice synthesize histamine.     

Experiences with the application of QSAR in the routine of the notification procedure

IL-8 is a recently described chemokine that increases polymorphonuclear neutrophil infiltration and has been implicated in inflammatory pathology. This study assesses monocyte (M phi) interleukin-8 (IL-8) levels in severe trauma patients (injury severity score > 16) who have elevated levels of M phi cell-associated tumor necrosis factor alpha (TNF alpha), a major marker for systemic inflammatory response syndrome after injury. We demonstrate elevated (p = .0007) levels of M phi IL-8 only in those trauma patients who also have increased (p = .0001) M phi-secreted TNF alpha whereas the patients having normal M phi-secreted TNF alpha levels have normal or even decreased M phi IL-8 production. There is no association between M phi IL-8 production and cell-associated TNF alpha levels. M phi induction by Fc gamma RI cross-linking, a common induction pathway in trauma patients' M phi that increases the production of both cell-associated and secreted TNF alpha, can also increase (p = .0022) M phi IL-8 levels. Therefore, post-trauma elevation of M phi IL-8 levels may be associated with increased secreted TNF alpha resulting from, at least in part, Fc gamma RI cross-linking stimulation in vivo.     

Nitric oxide production in mouse peritoneal macrophages enhanced with glycyrrhizin

The enhancement of nitric oxide (NO) production in glycyrrhizin (GL)-induced macrophages (M phi) in response to lipopolysaccharide (LPS) was investigated. No production in GL-induced macrophage culture supernatants was stimulated in response to LPS (10 micrograms/ml) for 24- or 48- h cultures, and these levels were compared three times with the levels in saline-induced peritoneal exudate cell cultures. Furthermore, M phi induced with proteose peptone (PP) containing GL could generate greater NO production than M phi induced with PP alone. However, no stimulation of NO production was observed by addition of GL in the cultures of M phi induced with thioglycollate or Bacillus Calmette Guerin. Moreover, GL-induced M phi showed cytostasis against such tumor target cells as L 1210 and P 388 lymphoma cell lines. These observations indicate that GL can activate the M phi in vivo system and stimulate NO production in response to LPS.     

Increase in histamine synthesis by liver macrophages in CCl4-injured mast cell-deficient W/Wv mice

This study set out to examine the possible role of liver macrophages in histamine synthesis in the injured liver. The effects of the hepatotoxins Escherichia coli lipopolysaccharide (LPS) and CCl4 on histamine synthesis in the liver of mice were evaluated. C3H/HeJ mice were resistant to LPS in including histidine decarboxylase (HDC) in the liver compared with C3H/HeN mice and mast cell-deficient W/Wv mice. However, C3H/HeJ mice did respond strongly to another hepatotoxin, CCl4, leading to a significant increase in HDC activity. CCl4 also caused a marked increase in HDC activity and histamine levels in the liver of W/Wv mice. In addition, injection of CCl4 produced a large increase in the activity of HDC in the spleen and lung of W/Wv mice. HDC activity was confined to the nonparenchymal cells, with parenchymal cells expressing essentially no HDC activity. The CCl4-induced increase in HDC activity was confined, at least in part, to the liver macrophages. These results indicate that the macrophages are responsible for the increase in HDC-dependent histamine production in the liver caused by the injection of hepatotoxins. The possible role of histamine in liver regeneration after injury is discussed.     

Tumorigenic conversion of a non-tumorigenic rat urothelial cell line by overexpression of H2O2-generating peroxisomal fatty acyl-CoA oxidase

Two of the major cutaneous consequences of ultraviolet (UV) radiation exposure are immunosuppression and the development of skin cancer. This study examined whether these effects are genetically determined. Suppression of contact hypersensitivity by local, low-dose UV radiation was examined in what have been termed "UV-susceptible" and "UV-resistant" strains of mice. C3H/HeJ mice ("UV resistant") were resistant to the adverse effects of low-dose UV radiation when normal doses of hapten were applied to UV-irradiated skin; however, they were sensitive when the amount of hapten used for sensitization was reduced. A similar effect was observed in BALB/c mice ("UV resistant") and when the hapten was dimethylbenz(a)anthracene, thus indicating that the genetic variation was not strain or hapten specific. Despite the fact that some strains were sensitive and some were resistant to low-dose UV radiation when high doses of hapten were employed, all strains initially sensitized to hapten through UV-irradiated skin were found to be unresponsive when rechallenged on normal skin, no matter what the initial sensitizing dose of hapten was. To determine whether other biologic effects of UV also exhibited genetic variation, C3H/HeN and C3H/HeJ mice were compared for susceptibility to UVB-induced skin cancer formation. C3H/HeJ mice developed significantly more tumors than C3H/HeN mice when subjected to a single dose of UV radiation followed by repeated exposure to the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate. These studies provide strong evidence that genetic factors influence individual susceptibility to the biologic effects of UV radiation.