Analysis of the NUDT15 gene and metabolites of azathioprine in Japanese patients with inflammatory bowel disease.

BACKGROUND: Thiopurines continue to play an important role in the treatment of inflammatory bowel disease (IBD). It is well known that thiopurines can cause several adverse reactions. Especially, hematopoietic toxicity may lead to severe agranulocytosis. In a previous prospective study, we investigated the relationship between inosine triphosphate pyrophosphatase (ITPA) c.94c > a polymorphism, 6-thioguanine nucleotide (6-TGN) concentration and toxicity. METHODS: To clarify the cause of thiopurine toxicity, we analysed nucleoside disphosphate-linked moiety X-type motif 15 (NUDT15) gene polymorphisms, i.e., R139C, V18I, and V19_V19insGV, and measured 6-mercaptopurines and 6-methylmercaptopurines (6-MMP) using the archived blood samples collected from 49 IBD patients for our previous study. RESULTS: The ITPA c.94c > a polymorphism was detected in 19 patients (38.7%, all heterozygous). The R139C polymorphism was found in 10 patients (20.4%, 1 homozygous, 9 heterozygous), V18_V19insGV in 7 patients (14.3%, all heterozygous), and V18I in 2 patients (4.08%, all heterozygous). Although R139C was more strongly associated with leukopenia than c.94c > a, there were no significant correlations with 6-TGN and 6-MMP levels, as for c.94c > a. The leukopenia incidence rates for each gene polymorphism were 0% in those with all wild-type genes, 21.4% for c.94c > a only, 42.9% for NUDT15 polymorphism (s) only, and 80.0% for both polymorphisms. CONCLUSIONS: All cases of leukopenia were associated with ITPA c.94c > a and/or polymorphism of NUDT15 and the risk of developing leukopenia was synergistically increased by ITPA and NUDT15 gene polymorphism. However, there was no association between the level of azathioprine metabolites and these polymorphisms.

Puromycin-sensitive aminopeptidase is required for C2C12 myoblast proliferation and differentiation.

The ubiquitin-proteasome system is a major protein degradation pathway in the cell. Proteasomes produce several peptides that are rapidly degraded to free amino acids by intracellular aminopeptidases. Our previous studies reported that proteolysis via proteasomes and aminopeptidases is required for myoblast proliferation and differentiation. However, the role of intracellular aminopeptidases in myoblast proliferation and differentiation had not been clarified. In this study, we investigated the effects of puromycin-sensitive aminopeptidase (PSA) on C2C12 myoblast proliferation and differentiation by knocking down PSA. Aminopeptidase enzymatic activity was reduced in PSA-knockdown myoblasts. Knockdown of PSA induced impaired cell cycle progression in C2C12 myoblasts and accumulation of cells at the G2/M phase. Additionally, after the induction of myogenic differentiation in PSA-knockdown myoblasts, multinucleated circular-shaped myotubes with impaired cell polarity were frequently identified. Cell division cycle 42 (CDC42) knockdown in myoblasts resulted in a loss of cell polarity and the formation of multinucleated circular-shaped myotubes, which were similar to PSA-knockdown myoblasts. These data suggest that PSA is required for the proliferation of myoblasts in the growth phase and for the determination of cell polarity and elongation of myotubes in the differentiation phase.

Fatty Acids as Useful Serological Markers for Crohn's Disease.

BACKGROUND: We have previously reported that patients with Crohn's disease (CD) have a very specific erythrocyte membrane phospholipid fatty acid profile. The findings of this study suggest that the activities of enzymes involved in the metabolism of linoleic acid (LA), that is, delta-6 desaturase, are higher in CD patients than in healthy individuals. METHODS: We evaluated the utilities of various fatty acid compositions of the plasma (p-) as new serological markers for CD compared to those of erythrocyte membranes (e-). RESULTS: Fifty CD patients and 50 healthy individuals were enrolled. In both plasma and erythrocyte membranes, the weight percentages of palmitic acid (PA) were significantly higher, while those of LA were significantly lower in CD patients than in controls. Fatty acids with high sensitivity and specificity were p-PA (0.86 and 0.74) and e-PA (0.80 and 0.74). With PA and LA as a CD fatty acid index (CDFAi), that is, CDFAi = (PA/LA), the sensitivity and specificity of plasma CDFAi (p-CDFAi) and e-CDFAi were 0.80 and 0.80; and 0.82 and 0.88 respectively. CONCLUSION: In CD patients, various fatty acids were specifically altered in both plasma and erythrocytes, and p-PA and p-CDFAi are potentially useful as new serological markers for CD.

The 1st step initiation essential for allergen-specific IgE antibody production upon the 2nd step: Induction of non-specific IgE+ small B cells containing secondly-sensitized allergen-specific ones in mice firstly-sensitized with an allergen.

There was a significant amount of non-specific, but not of allergen (e.g., papain, mite feces and four kinds of pollen)-specific, IgE antibodies (Abs) in the sera of normal mice. An i.n. injection of each allergen without adjuvant into mice caused an increase in total IgE Ab titers with a similar time course in the serum. However, the stage of initiation of allergy varied from allergen to allergen. Submandibular lymph node cells from normal mice contained papain-, but not mite feces- or pollen-specific IgE+ cells and an i.n. injection of papain induced papain-specific IgE Abs in the serum. In contrast, one (i.n.) or two (i.n. and s.c) injections of mite feces induced neither mite feces-specific IgE+ cells in the lymph nodes nor mite feces-specific IgE Abs in the serum. I.n. sensitization with cedar pollen induced cedar pollen-specific IgE+ small B cells in the lymph nodes on Day 10, when non-specific IgE Ab titers reached a peak in the serum, implying induction of related allergen-specific IgE+ small cells as well. In fact, a second (s.c.) injection of ragweed (or cedar) pollen into mice sensitized i.n. once with cedar (or ragweed) pollen, but not with mite feces, induced a large amount of ragweed (or cedar) pollen-specific IgE Abs in the serum. These results indicate that when firstly-sensitized non-specific IgE+ small B cells in mouse lymph nodes include some secondly-sensitized allergen-specific ones, mice produce IgE Abs specific for the secondly-injected allergen.

Neutropenia related to an azathioprine metabolic disorder induced by an inosine triphosphate pyrophosphohydrolase (ITPA) gene mutation in a patient with PR3-ANCA-positive microscopic polyangiitis
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A 68-year-old Japanese man was monitored for chronic kidney disease (CKD), with unknown primary disease starting in 2014. His serum creatinine (sCr) was stable at ~ 2.5 mg/dL for ~ 2 years. Two weeks before admission, he had bloody sputum, and sCr increased to 4.63 mg/dL. Soon after admission, the patient developed a high fever with pigment spots on the legs. A kidney biopsy was performed. The kidney specimens showed necrotizing and crescentic glomerulonephritis without granuloma formation. An additional blood-sampling test revealed high titers of PR3-ANCA, and we diagnosed PR3-ANCA-positive microscopic polyangiitis (MPA). Treatment with intravenous steroid pulse therapy and intermittent pulse intravenous cyclophosphamide therapy was started for remission induction. With these treatments, sCr improved to ~ 3.0 mg/dL. Azathioprine (AZA) was added for remission-maintenance therapy. Three days later, the dose of AZA was increased from 50 to 100 mg/day, and the number of neutrophils decreased to 30/µL. After withdrawal of AZA, neutrophil levels gradually recovered. We suspected that an abnormal metabolism of AZA was responsible for the neutropenia. Therefore, we analyzed three AZA metabolism-associated genes for mutations: thiopurine S-methyltransferase (TPMT), inosine triphosphate pyrophosphohydrolase (ITPA), and nucleoside diphosphate linked moiety X-type motif 15 (NUDT15), and we identified ITPA 94C>A mutation. This was a rare case of PR3-positive MPA with AZA-induced severe neutropenia that was possibly due to an ITPA gene mutation. This case suggests that ITPA gene mutation is related to the adverse reactions of AZA in Japanese patients. We have to pay attention to severe neutropenia when we use AZA, especially in Asian patients with CKD.
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Alarm pheromone is detected by the vomeronasal organ in male rats.

It is widely known that a stressed animal releases specific pheromones, possibly for alarming nearby conspecifics. We previously investigated an alarm pheromone in male rats and found that this alarm pheromone evokes several responses, including increases in the defensive and risk assessment behaviors in a modified open-field test, and enhancement of the acoustic startle reflex. However, the role of the vomeronasal organ in these pheromone effects remains unclear. To clarify this point, vomeronasal organ-excising or sham surgeries were performed in male rats for use in 2 experimental models, after which they were exposed to alarm pheromone. We found that the vomeronasal organ-excising surgery blocked the effects of this alarm pheromone in both the modified open-field test and acoustic startle reflex test. In addition, the results of habituation/dishabituation test and soybean agglutinin binding to the accessory olfactory bulb suggested that the vomeronasal organ-excising surgery completely ablated the vomeronasal organ while preserving the functioning of the main olfactory system. From the above results, we showed that the vomeronasal organ plays an important role in alarm pheromone effects in the modified open-field test and acoustic startle reflex test.

Role of cytokines in lavage or drainage fluid after hemithyroidectomy in wound healing: involvement of histamine in the acceleration and delay of wound healing.

Wound healing is a sophisticated biologic process. In the case of hemithyroidectomy, the operation time is relatively short with small tissue damage and without skin excision, and bacterial contamination before, during, and after the operation is uncommon. Here, we explored which cytokine(s) affected the rates of healing of skin wounds after hemithyroidectomy of 29 patients. We assessed the amounts of cytokines (e.g., interleukin-6, platelet-derived growth factor, basic fibroblast growth factor, vascular endothelial growth factor, and tumor necrosis factor-α) in either the preoperative or postoperative lavage fluids, or in the drainage fluids on postoperative days (PODs) 1-8. All of these cytokines showed a similar pattern; after reaching a peak on POD1, the production fell sharply on POD2-8, revealing that wound healing commenced on POD1. The rates of wound healing were inversely related to the levels of histamine in six patients (i.e., those with the three largest and those with the three smallest total volumes of drainage fluid on POD1): high (or low) levels of histamine in the postoperative lavage fluids with low (or high) levels in the drainage fluids on POD1 caused earlier (or the delay of) wound healing, suggesting involvement of histamine in the acceleration and delay of wound healing.

Essential role of macrophages in the initiation of allergic rhinitis in mice sensitized intranasally once with cedar pollen: regulation of class switching of immunoglobulin in B cells by controlling interleukin-4 production in T cells of submandibular lymph nodes.

The production of allergen-specific IgE antibodies (Abs) in allergen-sensitized patients or animals has a mutual relationship with the immunologic response leading to allergic rhinitis. We recently reported that, after an intranasal injection of cedar pollen into mice, an interleukin-4 (IL-4)-dependent increase in serum nonspecific IgE Abs was a prerequisite for the production of serum allergen-specific IgE Abs. Here, we explored which lymphoid organs were responsive to the intranasally injected allergen and how IL-4 and IgE Abs were produced in the lymphocytes. Time-dependent changes in the total cell numbers and in in vitro IgE Ab production in various lymphoid organs revealed that the submandibular lymph nodes were the main responsible organ. After treatment with allergen (for IgE production) or allergen and complete Freund's adjuvant (for IgG production), we separated submandibular lymph node cells into macrophage-, lymphocyte-, and granulocyte-rich populations by discontinuous Percoll density-gradient centrifugation. Unexpectedly, bulk cells, but not the lymphocyte- or macrophage-rich populations, produced significant amounts of IL-4, IgE, and IgG; whereas production was restored by addition of Mac-1(+) cells from the macrophage-rich to the lymphocyte-rich fraction. Furthermore, a combination of the lymphocyte-rich population (for IgG [or IgE]) production) and the macrophage-rich population (for IgE [or IgG]) production) produced a large amount of IgE (or IgG). These results indicate that, in the initiation of allergic rhinitis, macrophages in the submandibular lymph nodes are essential not only for IL-4 or immunoglobulin production, but also for class switching of immunoglobulin in lymphocytes.

Regulation of epithelial sodium transport via epithelial Na+ channel.

Renal epithelial Na+ transport plays an important role in homeostasis of our body fluid content and blood pressure. Further, the Na+ transport in alveolar epithelial cells essentially controls the amount of alveolar fluid that should be kept at an appropriate level for normal gas exchange. The epithelial Na+ transport is generally mediated through two steps: (1) the entry step of Na+ via epithelial Na+ channel (ENaC) at the apical membrane and (2) the extrusion step of Na+ via the Na+, K+-ATPase at the basolateral membrane. In general, the Na+ entry via ENaC is the rate-limiting step. Therefore, the regulation of ENaC plays an essential role in control of blood pressure and normal gas exchange. In this paper, we discuss two major factors in ENaC regulation: (1) activity of individual ENaC and (2) number of ENaC located at the apical membrane.

Spontaneous rejection of intradermally transplanted non-engineered tumor cells by neutrophils and macrophages from syngeneic strains of mice.

It is not surprising that tumors arising spontaneously are rarely rejected by T cells, because in general they lack molecules to elicit a primary T-cell response. In fact, cytokine-engineered tumors can induce granulocyte infiltration leading to tumor rejection. In the present study, we i.d. injected seven kinds of non-engineered tumor cells into syngeneic strains of mice. Three of them (i.e. B16, KLN205, and 3LL cells) continued to grow, whereas four of them (i.e. Meth A, I-10, CL-S1, and FM3A cells) were spontaneously rejected after transient growth or without growth. In contrast to the i.d. injection of B16 cells into C57BL/6 mice, which induces infiltration of TAMs into the tumors, the i.d. injection of Meth A cells into BALB/c mice induced the invasion of cytotoxic inflammatory cells, but not of TAMs, into or around the tumors leading to an IFN-γ-dependent rejection. On day 5, the cytotoxic activity against the tumor cells reached a peak; and the effector cells were found to be neutrophils and macrophages. The i.d. Meth A or I-10 cell-immunized, but not non-immunized, mice rejected i.p.- or i.m.-transplanted Meth A or I-10 cells without growth, respectively. The main effector cells were CTLs; and there was no cross-sensitization between these two kinds of tumor cells, suggesting specific rejection of tumor cells by CTLs from i.d. immunized mice. These results indicate that infiltration of cytotoxic myeloid cells (i.e. neutrophils and macrophages, but not TAMs) into or around tumors is essential for their IFN-γ-dependent spontaneous rejection.

Transgene number-dependent, gene expression rate-independent rejection of Dd -, Kd-, or Dd Kd -transgened mouse skin or tumor cells from C57BL/6 Db Kb mice.

Recipient cells migrating into the transplantation site of an allograft recognize histocompatibility antigens on the grafts and are cytotoxic against the grafts. Although the alloreactive immune response is predominantly directed at the major histocompatibility complex (major histocompatibility complex [MHC]; H-2 in mice) class I molecules, the basic mechanisms of allograft rejection (e.g., ligand-receptor interaction) remain unclear, because of the polymorphism and complexity of the MHC. To examine the role of MHC class I molecules in allograft rejection, D(d) , K(d) or D(d) K(d) -transgenic skin or tumor cells we established on a C57BL/6 (D(b) K(b) ) background and transplanted into C57BL/6 mice. Skin grafts from allogeneic (i.e., BALB/c, B10.D2, and BDF1) strains of mice were rejected from C57BL/6 mice on days 12-14 after grafting, whereas isografts were tolerated by these mice. Unexpectedly, skin grafts from D(d) -, K(d) -, and D(d) K(d) -transgenic C57BL/6 mice were rejected on days 12-14 in a transgene expression rate-independent manner from 9/19 (47%), 20/39 (51%), and 12/17 (71%) of C57BL/6 mice, respectively. Similarly, intradermally transplanted allogeneic (i.e., Meth A), but not syngeneic (i.e., EL-4), tumor cells were rejected from C57BL/6 mice; the growth of D(d) - or K(d) -transfected EL-4 cells was delayed by 10-13 days; and 4/10 (40%) of D(d) K(d) -transfected tumor cells were rejected from C57BL/6 mice. These results indicate that D(d) and K(d) genes are equivalent as allogeneic MHC class I genes and that C57BL/6 (D(b) K(b) ) mice reject D(d) -, K(d) -, or D(d) K(d) -transgened skin or tumor cells in a transgene number-dependent, gene expression rate-independent manner.

Monte Carlo study of dosimetric impact of gadolinium contrast medium in transverse field MR-Linac system.

The aim of this study is to evaluate the dosimetric impact of gadolinium contrast medium (Gadovist) in a transverse MR-Linac system using Monte Carlo methods. The dose distributions were calculated using two heterogeneous multi-layer phantoms consisting of Gadovist, water, bone, and lung. The photon beam was irradiated with a filed size of 5 × 5 cm2, and a transverse magnetic field of 0-3.0 T was applied perpendicular to the incident photon beam. Next, dose distributions for brain, head and neck (H&N), and lung cancer patients were calculated using a patient voxel-based phantom with and without replacing the patient's GTV with Gadovist. The dose at the water-Gadovist interface increased by 8% without a magnetic field. A similar dose increment was observed at 0.35 T. In contrast, the dose increment at the water-Gadovist interface was small at 1.5 T and a dose decrement of 5% was observed at 3.0 T. The dose variation at the lung-Gadovist interface was larger than that at the water-Gadovist interface. The mass collision stopping power ratio for Gadovist was 7% lower than that for water, whereas, the electron fluence spectra at the water-Gadovist interface increased by 17.5%. In a patient study, Gadovist increased the Dmean for brain, H&N, and lung cancer patients by 0.65-8.9%. The dose variation due to Gadovist grew large in the low-dose region in H&N and lung cancer. The GTV dose variation due to Gadovist in all treatment site was below 2% at 0-3 T if the Gadovist concentration was lower than 0.2 mmol/ml-1.

Transplantation of Graft Anti-Host Cytotoxic T Lymphocytes Along with Allogeneic Bone Marrow Skips Macrophage-Induced Graft-Versus-Host Disease.

Graft-versus-host disease (GVHD) is a physiological response of the graft to allogeneic hosts. However, the effector cells, affected organ(s), and cytokines in the GVHD remain controversially discussed, without having determined a particular cytotoxic activity of the graft against the host. After i.v. injection of C57BL/6 (H-2b) spleen cells into irradiated BDF1 (H-2b/d) mice, the hosts developed interferon-gamma (IFN-γ)-dependent bone marrow (BM) GVHD on days 5-17. When H-2DdKd transgenic H-2b lymphoma cells were i.p. inoculated into irradiated, H-2b splenocyte-transplanted H-2b/d mice, the infiltration of macrophages cytotoxic against H-2DdKd transgenic H-2b mouse skin epithelia (a GVHD activity) into the peritoneal cavity preceded several days the infiltration of interleukin (IL)-2-dependent cytotoxic T lymphocytes (CTLs) to achieve a graft-versus-leukemia (GVL) effect. In contrast, allogeneic BM transplanted alone into the irradiated mice did not induce GVHD for 44 days, whereas i.v. injection of graft anti-host macrophages or graft anti-host CTLs along with allogeneic BM, respectively, induced GVHD or promoted the GVL effect in the absence of GVHD. These results revealed that macrophage-induced GVHD and the CTL-mediated GVL effect were a set (Th1: IFN-γ/IL-2) response of the graft to allogeneic hosts and leukemia cells, respectively, and that graft T cell activation rather than inhibition skipped GVHD after BM transplantation.

Impact of the cavity on sinus wall dose in magnetic resonance image-guided radiation therapy.

PURPOSE: This study aims to investigate the impact of the cavity on the sinus wall dose by comparing dose distributions with and without the sinus under magnetic fields using Monte Carlo calculations. METHODS: A water phantom containing a sinus cavity (Empty) was created, and dose distributions were calculated for 1, 2, and 4 irradiation fields with 6 MV photons. The sinus in the phantom was then filled with water (Full), and the dose distributions were calculated again. The sinus was set to cubes of 2 cm and 4 cm. The magnetic field was applied to the transverse and inline direction under the magnetic flux densities of 0 T, 0.35 T, 0.5 T, 1.0 T, and 1.5 T. The dose distributions were analyzed by the dose difference, dose volume histogram, and D2 with sinus wall thicknesses of 1 and 5 mm. RESULTS: D2 in the "Empty" sinus wall under transverse magnetic fields for the 1-field and 4-field cases was 51.9% higher and 3.7% lower than that in the "Full" sinus wall at 1.5 T, respectively. Meanwhile, D2 in the Empty sinus wall under inline magnetic fields for 1-field and 4-fields was 2.3% and 2.6% lower than that in the "Full" sinus at B = 0 T, respectively, whereas D2 was 0.9% and 0.7% larger at 1.0 T, respectively. CONCLUSIONS: The impact of the cavity on the sinus wall dose depends on the magnetic flux density, direction of the magnetic field and irradiation beam, and number of irradiation fields.

Comparison of dose distributions between transverse magnetic fields of 0.35 T and 1.5 T for radiotherapy in lung tumor using Monte Carlo calculation.

We investigated the impact of the transverse magnetic fields of 0.35 T and 1.5 T on the dose distributions for a 6 MV beam, by using a thorax phantom with a lung tumor. First, the dose distributions in the magnetic flux densities of 0 T, 0.35 T, and 1.5 T were compared by increasing the number of irradiation fields. Next, the dose distributions for stereotactic body radiotherapy (SBRT) with 5-fields for an isolated lung tumor was compared in transverse magnetic fields. All dose distributions were calculated by the Monte Carlo method. The prescription doses for SBRT with 5-fields was 48 Gy for D95 (dose covering 95% volume) in the planning target volume (PTV). The dose distributions were analyzed by the dose difference map (DD map), dose volume histogram (DVH), and dose indices. For the 1-field, the dose distributions were more affected at 1.5 T rather than 0.35 T. The DVHs for PTV at 1.5 T almost agreed with those at 0 T for more than 5-fields. In contrast, the D98 in the PTV at 0.35 T reduced constantly by 6.0% with more than 5-fields. The D95 in PTV for SBRT with 5-fields was 9.0% lower at 0.35 T and 2.5% higher at 1.5 T, in comparison with that at 0 T. For dispersed irradiation angles of more than 5-fields, it is more desirable to use the magnetic flux density of 1.5 T than 0.35 T for the radiotherapy in the lung tumor.

Impact of lung density on isolated lung tumor dose in VMAT using inline MR-Linac.

PURPOSE: This study investigated the impact of lung density on the isolated lung tumor dose for volumetric modulated arc therapy (VMAT) in an inline magnetic resonance linear accelerator (MR-Linac) using the Monte Carlo (MC) simulation. METHODS: CT images of the thorax phantoms with lung tumors of 1, 2, and 3 cm diameters were converted into voxel-base phantoms with lung densities of 0.1, 0.2, and 0.3 g/cm3, respectively. The dose distributions were calculated for partial-arc VMAT. The dose distributions were compared using dose differences, dose volume histograms, and dose volume indices. RESULTS: In all cases, the inline magnetic field significantly enhanced the lung tumor dose compared to that at 0 T. For the 1 cm lung tumor, the inline magnetic field of 1 T increased the minimum dose of 95% of the Planning target volume (PTV D95) by 14.0% in 0.1 g/cm3 lung density as compared to that in 0.3 g/cm3 at 0 T. In contrast, at 0 and 0.5 T, the PTV D95 in 0.3 g/cm3 lung density was larger than that in lung density of 0.1 g/cm3. For the 2 cm lung tumor, a similar tendency to 1 cm was observed, whereas the dose impact of lung density was smaller than that for 1 cm. For the 3 cm lung tumor, the lung tumor dose was independent of lung density at 0.5 T and 1.0 T. CONCLUSION: The inline MR-Linac with the magnetic field over 1 T can enhance the PTV D95 for VMAT regardless of the lung density.

IFN-γ Control of an Effector/Target Combination for Skin Allograft Rejection: Macrophage/Skin Components in Normal Mice or T Cell/Endothelial Cells in IFN-γ-Deficient Mice.

Organ, skin, or cell allografts are acutely rejected from normal mice, whereas vascularized organ allografts, but not allografted Meth A cells, are rejected from interferon-γ (IFN-γ)-deficient mice. Here we explored effector/target combinations for i.p. allografted Meth A (cytotoxic T lymphocyte [CTL]-resistant) or RLmale1 (CTL-susceptible) cells into or for BALB/c skin (skin components: CTL resistant) onto normal or IFN-γ-deficient C57BL/6 mice. After allografting, normal mice showed more infiltration but only a little thrombosis/hemorrhage. Monocyte/macrophage MHC receptor (MMR)+ macrophages (on days 5-10) and T cell receptor (TCR)+ CTLs (on days 7-9) were cytotoxic against Meth A cells or skin components and RLmale1 cells, respectively, and the allografts were rejected. After allografting into IFN-γ-deficient mice, MMR- macrophages and highly activated TCR+ CTLs were induced, and the mice died of hemorrhagic ascites with Meth A cells and more acutely rejected RLmale1 cells. The CTLs on days 4-6 were inactive toward skin components at an in vivo effector/target ratio but injured endothelial cells to cause severe thrombosis/hemorrhage and more acute rejection of skin allografts. These results indicate that IFN-γ-dependent MMR expression was essential for macrophage-mediated cytolysis of allogeneic skin components and that IFN-γ-deficient mice more acutely rejected skin allograft by causing CTL-induced injury to endothelial cells.

Rejection of intradermally injected syngeneic tumor cells from mice by specific elimination of tumor-associated macrophages with liposome-encapsulated dichloromethylene diphosphonate, followed by induction of CD11b(+)/CCR3(-)/Gr-1(-) cells cytotoxic against the tumor cells.

Tumor cell expansion relies on nutrient supply, and oxygen limitation is central in controlling neovascularization and tumor spread. Monocytes infiltrate into tumors from the circulation along defined chemotactic gradients, differentiate into tumor-associated macrophages (TAMs), and then accumulate in the hypoxic areas. Elevated TAM density in some regions or overall TAM numbers are correlated with increased tumor angiogenesis and a reduced host survival in the case of various types of tumors. To evaluate the role of TAMs in tumor growth, we here specifically eliminated TAMs by in vivo application of dichloromethylene diphosphonate (DMDP)-containing liposomes to mice bearing various types of tumors (e.g., B16 melanoma, KLN205 squamous cell carcinoma, and 3LL Lewis lung cancer), all of which grew in the dermis of syngeneic mouse skin. When DMDP-liposomes were injected into four spots to surround the tumor on day 0 or 5 after tumor injection and every third day thereafter, both the induction of TAMs and the tumor growth were suppressed in a dose-dependent and injection number-dependent manner; and unexpectedly, the tumor cells were rejected by 12 injections of three times-diluted DMDP-liposomes. The absence of TAMs in turn induced the invasion of inflammatory cells into or around the tumors; and the major population of effector cells cytotoxic against the target tumor cells were CD11b(+) monocytic macrophages, but not CCR3(+) eosinophils or Gr-1(+) neutrophils. These results indicate that both the absence of TAMs and invasion of CD11b(+) monocytic macrophages resulted in the tumor rejection.

Thiopurine S-methyltransferase and inosine triphosphate pyrophosphohydrolase genes in Japanese patients with inflammatory bowel disease in whom adverse drug reactions were induced by azathioprine/6-mercaptopurine treatment.

BACKGROUND: The main cause of azathioprine (AZA)/6-mercaptopurine (6MP)-induced adverse reactions is a reduction in the activities of the metabolizing enzymes thiopurine S-methyltransferase (TPMT) and inosine triphosphate pyrophosphohydrolase (ITPA). Adverse reactions develop at a high frequency in Japanese patients at half the dose required for European and American patients; however, the association with TPMT and ITPA gene polymorphisms in Japanese has not been fully investigated. METHODS: Gene mutations of TPMT and ITPA, the major AZA/6-MP -metabolizing enzymes, were investigated retrospectively in 16 Japanese patients with inflammatory bowel disease (IBD) in whom AZA/6MP treatment induced adverse reactions. RESULTS: The TPMT gene was found to have a wild-type sequence in all patients, but in the ITPA gene a mutation, 94C>A, was detected at a rate of 50% (8/16), with 83.3% (5/6) occurring in patients with acute bone marrow suppression and 75% (3/4) in those with agranulocytosis. The 94C>A allele frequency was 10 of 32 (0.313; 95% CI, 0.180-0.486). Adverse reactions developed earlier in patients with the 94C>A mutation. However, in half the patients, no gene polymorphism was noted. CONCLUSIONS: It is suggested that the ITPA gene mutation is closely related to the adverse reactions of AZA/6-MP in Japanese patients, and screening for the mutant allele is useful for predicting the most serious adverse reactions, agranulocytosis and acute bone marrow suppression.

Impact of inline magnetic fields on dose distributions for VMAT in lung tumor.

The purpose of this study is to investigate the impact of inline magnetic field on dose distribution for volumetric modulated arc therapy (VMAT) in lung tumors located at the chest wall and mediastinum. Two VMAT plans for a thorax phantom with lung tumors of 1 cm and 2 cm in diameter located at the chest wall were created by a treatment planning system. Next, five clinical VMAT plans for a non-small cell lung cancer (NSCLC) at early stages I and II of 5 cm or less in diameter were also used. The planning target volume (PTV) sizes were in the range from 11.1 to 82.7 cm3. The prescription dose was 60 Gy for D95 in the PTV. The VMAT dose distributions without and with uniform inline magnetic field of 0.5 T and 1.0 T were calculated using the Monte Carlo method. The dose distributions were analyzed by dose volume histograms, dose differences, and dose indices. In all VMAT plans, the PTV dose was enhanced by inline magnetic field. The dose enhancement was larger with 1.0 T than with 0.5 T. In phantom plans, D98 in the PTV with 0.5 T and 1.0 T increased by 2.9-6.6 Gy and 3.9-9.8 Gy, respectively, in comparison with that at 0 T. Similarly, in clinical plans, it increased by 2.2-6.0 Gy and 3.9-10.7 Gy, respectively. Thus, the VMAT with the inline magnetic field was proved useful for the dose enhancement in the lung tumor located at the chest wall and mediastinum.