Structure modifications based on KRN7000 and their SARs in activating NKT cells

α-Galactosylceramides, which can be recognized by natural killer T （NKT） cells, are now attracting more and more attention due to their therapeutic potential in cancer, infection and autoimmune diseases. Advances have been achieved in discovering compounds with better activities and efforts have been made to understand the structure-activity relationships （SARs） of these NKT cell ligands. In this review, we discuss the structure modifications based on KRN7000, the principal glycolipid used in the study of NKT cell stimulation, and the SARs based on these modified structures.

Hepatic stellate cells and innate immunity in alcoholic liver disease

Constant alcohol consumption is a major cause of chronic liver disease, and there has been a growing concern regarding the increased mortality rates worldwide. Alcoholic liver diseases (ALDs) range from mild to more severe conditions, such as steatosis, steatohepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma. The liver is enriched with innate immune cells (e.g. natural killer cells and Kupffer cells) and hepatic stellate cells (HSCs), and interestingly, emerging evidence suggests that innate immunity contributes to the development of ALDs (e.g. steatohepatitis and liver fibrosis). Indeed, HSCs play a crucial role in alcoholic steatosis via production of endocannabinoid and retinol metabolites. This review describes the roles of the innate immunity and HSCs in the pathogenesis of ALDs, and suggests therapeutic targets and strategies to assist in the reduction of ALD.

Epidemiology and gene markers of ulcerative colitis in the Chinese

Inflammatory bowel disease （IBD） includes two similar yet distinct conditions called ulcerative colitis （UC） and Crohn＇s disease （CD）. These diseases affect the digestive system and cause the inflammation of intestinal tissue, form sores and bleed easily. Most children with IBD are diagnosed in late childhood and adolescence. However, both UC and CD have been reported as early as in infancy. Most information pertaining to the epidemiology of IBD is based upon adult studies. Symptoms include abdominal pain, cramping, fatigue and diarrhea. Genetic factors play a significant role in determining IBD susceptibility. Epidemiological data support a genetic contribution to the pathogenesis of IBD. Recently, numerous new genes have been identified as being involved in the genetic susceptibility to IBD： TNF- 308A, CARD15 （NOD2）, MIF-173, N-acetyltransferase 2 （NAT2）, NKG2D （natural killer cell 2D）, STAT6 （signal transducer and activator of transcription 6）, CTLA-4 （cytotoxic T lymphocyte antigen-4）, MICA-MICB （major histocompatibility complex A and B）, HLA-DRB1, HLA class-Ⅱ, IL-18, IL-4, MICA-A5, CD14, TI R4, Fas-670, p53 and NF-kB. The characterization of these novel genes has the potential to identify therapeutic agents and aid clinical assessment of phenotype and prognosis in patients with IBD （UC and CD）.

Cytokines as critical co-stimulatory molecules in modulating the immune response of natural killer cells

Cytokines are involved in directing the activation of natural killer （NK） cells. NK cells are involved in the recognition of cells that have been altered; thus they do not recognize specific insults to the host, but when activated, are capable of destroying infected cells directly, as well as promoting the recruitment and response of the other components of the immune system by the release of cytokines and chemokines. It is these properties that have made NK cells a critical part of innate immunity and adaptive immunity, and they play a principal role linking innate and adaptive immunity by the recruitment of an adaptive immune response to an innate immune reaction.

Consumption of purple sweet potato leaves modulates human immune response: T-lymphocyte functions, lytic activity of natural killer cell and antibody production

AIM： To study the immunological effects of physiologica doses of purple sweet potato leaves （PSPL）. METHODS： The randomized crossover study （two periods, each lasting for 2 wk） involved 16 healthy non-smoking adults of normal weight. The 6-wk study consisted of a run-in （wk 1） PSPL diet （daily consumption of 200 g PSPL） or a control diet （low polyphenols, with the amount of carotenoids adjusted to the same level as that of PSPL） （wk 2-3）, washout diet （wk 4）, and switched diet （wk 5-6）. Fasting blood was collected weekly in the morning. T-lymphocyte function was assessed via the proliferation and secretion of immunoreactive cytokines. Salivary IgA secretion and the specific cytotoxic activities of cytotoxic T lymphocytes and natural killer （NK） cells were determined. RESULTS： The plasma 13-carotene level increased with time in both groups, while the plasma polyphenol level decreased in the control group, and no significant difference was detected between the two groups. Although p^asma polyphenol levels did not significantly increase in the PSPL group at the end of the study, they were significantly elevated in urine. PSPL consumption produced a significant increase in proliferation responsiveness of peripheral blood mononuclear cells （PBMC） and their secretion of immunoreactive IL-2 and IL-4. As well, lytic activity in NK cells was elevated in a time-dependent fashion. Salivary IgA secretion significantly decreased in control group after 2 wk, and returned to baseline following dietary switch to PSPL. CONCLUSION： Consumption of PSPL modulates various immune functions including increased proliferation responsiveness of PBMC, secretion of cytokines IL-2 and IL-4, and the lytic activity of NK cells. The responsible determinants of PSPL remain to be elucidated, as does the biological significance of the present observations.

Selective decrease in colonic CD56^+ T and CD161^+ T cells in the inflamed mucosa of patients with ulcerative colitis

AIM： To investigate the role of local colonic mucosal NK receptor-positive T （NKR＋ T） cells in the regulation of intestinal inflammation, we analyzed the population and function of these cells in ulcerative colitis （UC）. METHODS： Colonic mucosal tissues were obtained from colonoscopic biopsies of the descending colon from 96 patients with UC （51 endoscopically uninflamed, 45 inflamed） and 18 normal controls. Endoscopic appearance and histologic score at the biopsied site were determined by MaLts＇ classification. A single cell suspension was prepared from each biopsy by collagenase digestion. Two NKR^＋ T cell subsets, CD56^＋ （CD56^＋CD3^＋） T cells and CD161＋ （CD161^＋CD3^＋） T cells, were detected by flow cytometric analysis. Intracellular cytokine analysis for anti-inflammatory cytokine interleukin-10 （IL-10） was performed by in vitro stimulation with phorbol-myristateacetate （PMA） and ionomycin. RESULTS： CD56^＋ T cells and CD161^＋ T cells are present in the normal human colon and account for 6.7% and 21.3% of all mononuclear cells, respectively. The populations of both CD56＋ T cells and CD161^＋ T cells were decreased significantly in the inflamed mucosa of UC. In contrast, the frequency of conventional T cells （CD56 CD3^＋ cells and CD161CD3^＋ cells） was similar among the patient and control groups. The populations of NKR^＋ T cells were correlated inversely with the severity of inflammation, which was classified according to the endoscopic and histologic Marts＇ criteria. Interestingly, approximately 4% of mucosal NKR＋ T cells expressing IL-10 were detected by in vitro stimulation with PMA and ionomycin.CONCLUSION： Selective reduction in the population of colonic mucosal NKR＋T cells may contribute to the development of intestinal inflammation in UC.

Inositol hexaphosphate-induced enhancement of natural killer cell activity correlates with suppression of colon carcinogenesis in rats

AIM： To investigate the anti-neoplastic effect of inositol hexaphosphate （InsP6 or phytic acid） on dimethylhydrazine （DMH）-induced colon tumor in rats and its effect on blood natural killer （NK） cell activity. METHODS： Healthy Wistar rats, 4 wk old, were divided into control group （fed with common food） and InsP6 group （fed with common food＋2% sodium inositol hexaphosphate in the drinking water）, 15 rats in each group. Both groups were injected with 1,2-dimethylhydrazine subcutaneously （20 mg/kg body weight） once a week for 20 wk. Rats were killed after 21 wk. The whole large intestine was isolated to determine the general condition of tumors and to test blood NK cell activity by lactate-dehydrogenase-release assay. RESULTS： Administration of InsP6 significantly increased blood NK cell activity in DMH-induced colorectal tumor in rats. InsP6 group had a smaller tumor size on average and a smaller number of tumors than the control group. Its mortality was also higher than that in control. However, the variables of body weight and tumor incidence were not significantly different between the two groups. CONCLUSION： InsP6 can increase blood NK cell activity in DMH-induced colon tumor in rats and inhibit tumor growth and metastasis in rats.

Expression of prolactin receptor and response to prolactin stimulation of human NK cell lines

We have previously shown a critical role of prolactin (PRL) during maturation and anti-tumor effects of murine natural killer (NK) cells in vitro and in vivo. We extended that study by exploring the ability of human NK cell lines (NK-92 and YT cell) to express PRL receptor (PRL-R) and to respond to PRL stimulation in vitro. Both human NK cell lines constitutively expressed PRL-R on membrane and mRNA transcripts,NK-92 cells contained higher level of PRL-R than YT cells,which correlated to the enhanced capacity of the cells to proliferate and to lyse target cells in response to PRL stimulation in the presence of trace amount of IL-2 or IL-15 in vitro. Two differences between IL-2 and IL-15 in functioning on human NK cells were for the first time observed. PRL synergized with IL-15 to improve proliferation of NK cells in a dose-dependent manner without double peak manifesting like IL-2. Although PRL enhanced the cytotoxicity of IL-2 or IL- 15 activated NK cells,it exerted the function through up-regulating gene expression of perforin without influence of FasL in IL-2-stimulated NK cells,while in IL-15-stimulated NK cells,PRL did the function through up-regulating gene expression of both perforin and FasL but not IFNγ. PRL increased expressions of IL-2Rα on membrane and of IL-2 mRNA in cells,indicating that PRL up-regulated NK cell function by improving positive feedback between IL-2 and IL-2R. The similar results were also observed in network between IL-15 and IL-15R. These data indicate a potential role of PRL in human NK cell modulation.

Novel function of perforin in negatively regulating CD4^＋ T cell activation by affecting calcium signaling

Perforin is a pore-forming protein engaged mainly in mediating target T cell death and is employed by cytotoxic T lymphocytes （CTLs） and natural killer cells. However, whether it also plays a role in conventional CD4^＋ T cell function remains unclear. Here we report that in perforin-deficient （PKO） mice, CD4^＋ T cells are hyperproliferative in response to T cell receptor （TCR） stimulation. This feature of hyperproliferation is accompanied by the enhancement both in cell division and in IL-2 secretion. It seems that the perforin deficiency does not influence T cell development in thymus spleen and lymph node. In vivo, perforin deficiency results in increased antigen-specific T cell proliferation and antibody production. Furthermore, PKO mice are more susceptible to experimental autoimmune uveitis. To address the molecular mechanism, we found that after TCR stimulation, CD4^＋ T cells from PKO mice display an increased intracellular calcium flux and subsequently enhance activation of transcription factor NFAT1. Our results indicate that perforin plays a negative role in regulating CD4^＋ T cell activation and immune response by affecting TCR-dependent Ca^2＋ signaling.

Extranodal natural killer/T-cell lymphoma,nasal type:epidemiology study

Extranodal natural killer （NK）IT-cell lymphoma, nasal type （ENKTCL） is a rarely kind of non-Hodgkin lymphoma （NHL）. It is much more frequent in Asian and Latin American countries than other part of the world. It typically affects nasal cavity. In China, one of its endemically places, ENKTCL accounts for 74%-96% of nasal NHL. Patients with ENKTCL usually show a highly aggressive clinical course, and its etiology is unclear. However, it is already proved that ENKTCL is associated with Epstein-Barr virus （EBV） infection, regardless patients＇, ethnicity and areas. Some studies show that the risk will increase among several occupations, such as farmer, who are frequently exposure to pesticides and chemical solvent and risk can be cut down if taking some protective measures.

NKT cells in HIV-1 infection

Natural killer T （NKT） cells are a unique T cell population that have important immunoregulatory functions and have been shown to be involved in host immunity against a range of microorganisms. It also emerges that they might play a role in HIV-1 infection, and therefore be selectively depleted during the early stages of infection. Recent studies are reviewed regarding the dynamics of NKT depletion during HIV-1 infection and their recovery under highly active antiretroviral treatment （HAART）. Possible mechanisms for these changes are proposed based on the recent developments in HIV pathogenesis. Further discussions are focused on HIV＇s disruption of NKT activation by downregulating CDld expression on antigen presentation cells （APC）. HIV-1 protein Nefis found to play the major role by interrupting the intracellular trafficking of nascent and recycling CDld molecules.

Intrahepatic natural killer T cell populations are increased in human hepatic steatosis

AIM: To determine if natural killer T cell (NKT) populations are affected in nonalcoholic fatty liver disease (NAFLD). METHODS: Patients undergoing bariatric surgery underwent liver biopsy and blood sampling during surgery. The biopsy was assessed for steatosis and immunocyte infiltration. Intrahepatic lymphocytes (IHLs) were isolated from the remainder of the liver biopsy, and peripheral blood mononuclear cells (PBMCs) were isolated from the blood. Expression of surface proteins on both IHLs and PBMCs were quantified using flow cytometry. RESULTS: Twenty-seven subjects participated in thisstudy. Subjects with moderate or severe steatosis had a higher percentage of intrahepatic CD3+/CD56+ NKT cells (38.6%) than did patients with mild steatosis (24.1%, P = 0.05) or those without steatosis (21.5%, P = 0.03). Patients with moderate to severe steatosis also had a higher percentage of NKT cells in the blood (12.3%) as compared to patients with mild steatosis (2.5% P = 0.02) and those without steatosis (5.1%, P = 0.05). CONCLUSION: NKT cells are significantly increased in the liver and blood of patients with moderate to severe steatosis and support the role of NKT cells in NAFLD.

Reversing effects of traditional Chinese antitumor medicines on colorectal tumor immunosuppression of natural killer cell and T lymphocyte in vitro

Objective： Reversing effects of traditional Chinese medicines on colorectal tumor immunosuppressions of natural killer （NK） cell and T lymphocyte were analyzed to provide evidence on selecting medicines for patients according to the differ- ent types of tumor immunosuppression. Methods： Six traditional Chinese medicines, including Arsenious acid （AS）, Ligustra- zine hydrochloride （LHC）, Astragalus mongholicus bge （AMB）, Matrine N-oxide （MOX）, Polyporus umbellatus polysaccharide （PUPS） and Artesunate （ART）, were enrolled. The reversing effects on suppression of murine splenocyte transformation and NK killing activity were measured by 3-{4,5-dimethyl-2-thiazolyl}-2,5-diphenyl tetrazolium （MTT）, and the effects on the suppressed expression of intefieukin 2 receptor a （IL-2R（I）, CD3E＊~,＊ and CD3~.-~＊ were detected by flow cytometry （FCM）. The effects on immunosuppressive molecules were measured by enzyme linked immunosorbent assay （ELISA）, including transforming growth factor ~1 （TGF-~I）, vascular endothelial growth factor （VEGF）, interleukin 4 （IL-4）, IL-6, IL-10 and pros- taglandin （PG） E2. Results： （1） The reversing effects of AMB on the inhibition of NK killing and CD3 expression were the most significant; the effect of LHC on inhibition of CD3 expression was the strongest; the effects of AMB, PUPS and ART on inhibition of transformation were the greatest; and the effect of ART on inhibition of IL-2Ra expression was the strongest. （2） The correlated molecules of these medicines that exerted reversing effects on colorectal tumor immunosuppression were TGF-~I and IL-10. AMB had the highest down-regulating effect on the secretion of TGF-~I. AS and ART had the highest effects on IL-10. Conclusion： Reversing tumor immunosuppression through the down-regulation of immunosuppressive molecules is one of the novel antitumor mechanisms of traditional Chinese medicines. The clinical use of compounded prescriptions of ART combined with AMB and LHC should be considered to avoid the reduced treatment efficiency caused by tumor im- munosuppression.

EVALUATION OF THE ANTITUMOR ACTIVITY OF HUMAN IL－4 BY IN VITRO AND IN VIVO ASSAYS

The characteristics of rhuIL-4 induced cytotoxicity was detected in vitro by using ￣(51)Cr release assay and the anti-tumor activity of rhuIL-4 induced killer cell was evaluated in vivo by using a human tumor model in nude mice. huIL-4 can induce LAK activity from peripheral blood lymphocytes (PBMC) stimulated with phytohemagglutinin (PHA). Compared with the LAK activity induced by rhuIL-2, the cytotoxicity of the killer cells induced by rhuIL-4 to K562 and Raji cells was lower, but that to TBL-E, a human lymphoid leukernia cell line established in our laboratory, and PHA-activated blast cells (PHA-blasts) was of similar magnitude. In the cytotoxicity assay using PHA-blasts, the addition of PHA increased the IL-4-induced killer cell cytotoxicity by 131%, but had no effect on IL-2-induced killer cell cytotoxicity. This implies that IL-4 mainly induces CTL-like activity, while IL-2 mainly induces NK-like activity. An experimental human tumor model in nude mice was established by injection of TBL-E human leukemia cells. The anti-tumor activity of rhuIL-4 was evaluated by injection of human LAK cells induced from PHA-blasts by rhuIL-2+rhuIL-4 and human cytokines into tumor-bearing nude mice. The results showed that human LAK cells effectively inhibit the tumorigenicity of TBL-E cells in nude mice with an inhibition rate of 61 %. The antitumor effect of rhuIL-2 was better than that of rIL-4, and the antitumor effect of rhuIL-2+rhuIL-4 was similar to that of rhuIL-2, though the former delayed the occurence of tumors. Our data imply the potential application of human IL-4 in clinic, and provide an animal model to evaluate the anti-tumor activity of human cytokine(s) with species specificity.

Clinical significance of peripheral blood CD4^+ natural killer T cells in chronic hepatitis B virus infection

Objective:To understandtheclinicalsignificanceof CD4 + naturalkillerT(NK-T)cellsin chronic hepatitisB virus(HBV)infection.Methods :Peripheralbloodmononuclearcells(PBMCs)fromindividualswith chronicHBVinfectionwereseparated routinely.AfterInductionwithIL-12/IL-2for12d,theproportionof CD4 + NK-Tcellsin peripheralbloodwas determinedby fluorescenceactivatedcellsorter(FACS)analysis,andthe cytotoxicityof peripheralbloodlymphocytes(PBLs)was testedwitha4 h 51 Cr releaseassay.Results:After IL-12/IL-2induction,theproportionof CD4 + NK-Tcellswas(18.1±4.20)%, (6.95±2.85)%and(1.50±1.30)%inthe healthycontrol,CAHandAsCrespectively.Thatintheperipheralbloodof chronicHBVinfectedindividualswas lowerthanthatin thehealthycontrol.CD8+NK-Tcellswas(2.70±1.10)%, (2.20±1.40)%and(3.10±0.70)%respectively.In vitro cytotoxicityassaysagainstWishcellsrevealedthatthePBLscytotoxicityreducedin chronic HBVinfectedindividuals(P<0.05),andthatin AsCgroupwas significantlylowerin comparisonwithCHBand healthycontrolgroups.Thecytotoxicityof CD4 + NK-TcellsagainstWishcellscouldbe abolishedby treatingPBLs witheitheranti-CD4Abor anti-CD56Ab andcomplement,andpartiallydepleted by anti-CD8Ab.Conclusion:Theabnormalcellularimmunefunctionof chronicHBVinfectedindividualsmaybe associatedwiththedeficiency of CD4 + NK-Tcells.

The effect of Jinlong capsule on the immune function for intervened patients with primary liver cancer

Objective: The aim of this study was to evaluate the effect of Jinlong capsule on the immune function for intervened patients with primary liver cancer. Methods: Matched the inclusion criteria, 60 patients were selected and randomly divided into two groups. The treatment group had 30 cases treated with Jinlong capsule combined with the transcatheter arterial chemoembolization(TACE); the control group had 30 cases treated with TACE. Each group was treated 30 days as a cycle, which had completed at least two cycles. Indicators of cellular immune function about the activity of CD3, CD4, CD8, CD4/CD8 and natural killer(NK) cell were detected before and after treatment, then to compare and analysis with each other. Results: Before treatment, the activity of peripheral blood CD3, CD4, CD8, CD4/CD8 and NK cell in the two groups was no significant difference(P > 0.05); after treatment, the activity of CD3, CD4 and NK cell in the treatment group was significantly increased, the ratio of CD4/CD8 increased, and the value of CD8 decreased(P < 0.05), the activity of CD3, CD4 and NK cell in the control group was significantly decreased, the ratio of CD4/CD8 decreased(P < 0.05), and the value of CD8 slightly higher than before treatment(P > 0.05), the difference between the two groups indicated the statistical significance(P < 0.05). Incidence of gastrointestinal reactions, leucopenia, hemoglobin, platelet decline in the treatment group was lower than those in the control group, but without presenting the statistical significance(P > 0.05). Conclusion: Jinlong capsule with hepatic arterial infusion chemotherapy can improve the patients' immune function, and reduce the adverse reactions of interventional chemotherapy. Hence,it deserves to be promoted in clinically.

THE KINETICS OF CYTOPLASMIC GRANULE SECRETION IN NATURAL KILLER CYTOTOXICITY

Antisetum against purified cytoplasmic granules from rat LCL tumor cells. and protein A-gold immunoelectron microscopy were used to study the secretory events in lysis of YAC-1 tumor cells by rat LCL tumor cells or by isolated LGL from normal rats. After 30 min incubation of effector and target cells together, gold-labeled cytoplasmic granules were often seen concentrated in the area of the LGL adjacent to the bound YAC-1 target. Within 60 min, the granules were observed to move to the cell border near the conjugated site. At this point, the granules were fused with the cell membrane. and subsequently released the gold-labeled contents into the junction between the LGL and the target cell. Gold particles could be seen at the E-T interface, on the LGL surface, or sometimes on the target cell surface. These data provide direct evidence for the hypothesis that under conditions of active cytotoxicity, natural killer cells secrete their cytoplasmic granule contents leading to the deposition of granule material on the target cell surface and the eventual lysis of the cell.

EFFECT OF ACUPUNCTURE ON IL 2-IFN-NKC IMMUNOREGULATORY NETWORK IN MICE WITH TRANSPLANTED HEPATOCARCINOMA

: To study the effect of acupuncture on interleukin 2 (IL 2)-interferon(IFN)-natural killer cells (NKC) immunoregulatory network of mice with transplanted hepatocarcinoma(HAC) in order to provide new evidence for acupuncture treatment of hypofunction of immune system. Methods:The 28 HAC-vaccinated BALB/C mice are randomly divided into control group (n=14) and acupunctrue group (n=14). In the latter group bilateral'Dazhui' (BL 11) and 'Zusanili' (ST 36) which are located according to the same positions indicated in Comparative Anatomy of Macro-animals, are needled once every day, twelve sessions altogether. Twenty-four hours after the needling treatment the mice are killed and the spleen is taken out to be made into cell suspension for assaying concentrations of IL2 (MTT method) and NKC (colorimetric method) respectively. Obital Serum IFN is determined by using (CPE microplate staining), and the tumor mass is taken out and balanced with an analytical balance (1/10000) to calculate the tumor inhibition rate according to the formula. Results: The tumor weight of the mice of aupuncture group is obviously decreased (inhibition rate 43.06%) while the activity of the IL2 and NKC, and the IFN tiler are increased greatly, which have significant differences compared with those of control group (P>0.01). Conclusion:The results of the study show that acupuncture can strengthen the positive immunoregulatory function of the IL2 -IFN-NKC network in immune hypofunction mice bearing HAC.

A Mathematical Model of Breast Cancer and Mediated Immune System Interactions

Mathematical model that describes breast cancer and immune interactions were presented using system of differential equations to provide analytic and nnmeric framework of cancer-immune dynamics. Four types of immune cells-CTLs （cytotoxic T lymphocytes）, macrophages, NK （natural killer） and helper T cells-known to play the most significant roles in developing breast cancer immunity were modeled using differential equations. The model was then applied to different cancer growth rates and simulated using MATLAB software tool. The parameters of the model were based on experimental and clinical results from published articles. Results supported clinical studies that maximal breast cancer immunity depends mostly on each of the four immune cell types chosen. It was observed that for a given breast cancer growth rate, there was an optimal activation that maximized the response of the immune system. The effectiveness of the immune system resulted in the decrease in breast cancer killing rates. These results highlighted the importance of immune system activations in breast cancer development and treatment. Therefore, the model and its simulation provided a robust framework to better understand breast cancer progression and response to the immune system.

Inflammatory models drastically alter tumor growth and the immune microenvironment in hepatocellular carcinoma

Initiation and progression of hepatocellular carcinoma （HCC） is intimately associated with a chronically diseased liver tissue. This diseased liver tissue background is a drastically different microenvironment from the healthy liver, especially with regard to immune cell prevalence and presence of mediators of immune function. To better understand the consequences of liver disease on tumor growth and the interplay with its mi- croenvironment, we utilized two standard methods of fi- brosis induction and orthotopic implantation of tumors into the inflamed and fibrotic liver to mimic the liver condition in human HCC patients. Compared to non-diseased con- trols, tumor growth was significantly enhanced under fi- brotic conditions. The immune cells that infiltrated the tumors were also drastically different, with decreased numbers of natural killer cells but greatly increased num- bers of immune-suppressive CDllb^＋ Gr1^hi myeloid cells in both models of fibrosis. In addition, there were model- specific differences： Increased numbers of CD11b^＋ mye- loid cells and CD4^＋ CD25^＋ T cells were found in tumors in the bile duct ligation model but not in the carbon te- trachloride model. Induction of fibrosis altered the cytokine production of implanted tumor cells, which could have far- reaching consequences on the immune infiltrate and its functionality. Taken together, this work demonstrates that the combination of fibrosis induction with orthotopic tumor implantation results in a markedly different tumor mi- croenvironment and tumor growth kinetics, emphasizing the necessity for more accurate modeling of HCC pro- gression in mice, which takes into account the drastic changes in the tissue caused by chronic liver disease.