[Clinicopathological characteristics of NTRK-rearranged mesenchymal tumors in childhood].

Objective: To investigate the clinical and pathological features of pediatric NTRK-rearranged tumors. Methods: Four NTRK-rearranged soft tissue tumors and one renal tumor at Shanghai Children's Medical Center, Shanghai Jiaotong University and Singapore KK Women's and Children's Hospital from January 2017 to September 2019 were identified. Pan-TRK immunohistochemistry, and the ALK and ETV6 gene break-apart fluorescence in situ hybridizations (FISH) were performed. NTRK gene rearrangement was detected using sequencing-based methods. Results: There were 3 males and 2 females in this study. The patients were between 3 months and 13 years of age. Histologically, the tumors were infiltrative spindle cell tumors with variable accompanying inflammatory cells. Immunohistochemistry showed positive reactivity for pan-TRK in all tumors, with nuclear staining for NTRK3 fusion, and cytoplasmic staining for NTRK1 fusion. The molecular testing revealed NTRK gene fusions (one each of TPM3-NTRK1, ETV6-NTRK3 and DCTN1-NTRK1, and two cases of LMNA-NTRK1). Two patients were receiving larotrectinib. The others were are well without disease, with follow-up durations of 9 to 29 months. Conclusions: NTRK-rearranged mesenchymal tumors from soft tissue sites and kidney are identified. A novel DCTN1-NTRK1 fusion is described. Pan-TRK immunohistochemistry is useful for diagnosis. NTRK-targeted therapy may be an option for unresectable, recurrent or metastatic cases.

[Eosinophilic solid and cystic renal cell carcinoma with TSC2 gene mutations in children].

Objective: To study clinical pathological characteristics, immunohistochemical, molecular genetical changes and prognosis in pediatric eosinophilic solid and cystic renal cell carcinoma (ESC RCC) with TSC2 gene mutations. Methods: The tissue samples were collected from two pediatric ESC RCC patients between 2017 and 2018. The tissues were subjected to histological examination and immunohistochemistry using EnVision system. The TFE3, TFEB gene rearrangements were tested using FISH and molecular genetic study. The paraffin sections were used for DNA extraction, PCR amplification and NGS sequencing. Results: The two patients with ESC RCC were both male, aged at 9 years and 8 months, and 13 years, respectively. The tumors were from the right kidney, 5 cm and 7 cm in size, respectively, with solid and cystic changes in cross section, and grey-reddish or grey-whitish fish meat appearance. Microscopic observation revealed the tumors had fibrous capsules, which were infiltrated by the tumor cells. The tumor cells were diffusely distributed, round-shaped, or polygon-shaped, and had voluminous cytoplasm, eosinophilic cytoplasm, various sizes of vacuoles and clear cell-like appearance. There were papillary structures in some areas, with visible fiber septa. The nuclei were round and vesicular, with multi-nucleated cells and megakaryocytes. The mitoses were not seen. A few cystic structures were visible in different sizes, and capsule walls were covered with a single layer of spike-like tumor cells. Thick-walled blood vessels were seen in the stroma, with focal lymphocytic infiltration, eosinophilic necrosis, calcifications and cholesterol crystals. Immunohistochemistry of the tumor cells was positive for PAX8 (diffuse), CK20 (focal), CKpan (focal), CK10 (1 focal, 1 diffuse), INI1, vimentin, CD68, and Ki-67 (5%~10%); the tumor cells were negative for HMB45, S-100, Melan A, p53, desmin, TFE3, CK7, CK19, EMA, CD56, CgA, Syn, CD30, CD117, WT1 and SMA. Molecular genetic study showed that TFE3 and TFEB gene rearrangements were not detected by FISH. NGS sequencing showed TSC2 p.Lys574Ter (0.198) was found in patient one and TSC2 p.Arg406Ter (0.355) in patient two. Conclusions: ESC RCC in children is a rare disease, and can be misdiagnosed easily. It has unique pathological characteristics, and immunohistochemical, molecular and genetic changes. The prognosis is relatively good.

Cells with ganglionic differentiation frequently stain for VE1 antibody: a potential pitfall.

Mitogen-activated protein kinase (MAPK) pathway plays a major role in pediatric low-grade gliomas (pLGGs). Immunohistochemistry with mutant-specific antibody, VE1, has appeared to be the most affordable and rapidly deployable method to identify tumors with aberrant MAPK signaling pathway, by highlighting tumor with BRAFV600E mutation. Nonetheless, positive staining cases but not associated with BRAFV600E mutation are also seen. We analyzed 62 pLGGs for the two commonest genetic aberrations in MAPK pathway: KIAA1549-BRAF fusion, using reverse-transcriptase polymerase chain reaction, and BRAFV600E mutation, using VE1 antibody and Sanger sequencing. We recorded a specificity and accuracy rate of 68.75% and 75%, respectively, for VE1, when strong cytoplasmic staining is observed. Interestingly, we observed that cells with ganglionic features frequently bind VE1 but not associated with BRAFV600E mutation. Such observation was also confirmed in four cases of differentiating neuroblastoma. This false positive staining may serve as an important confounder in the interpretation of VE1 immunoreactivity with major therapeutic implication. It is important to confirm the presence of BRAFV600E mutation by DNA-based method, especially in tumor entities not known to, or rarely harbor such mutations.

C-Terminal Part of Glutamate-Ammonia-Ligase Adenyltransferase Gene Identified by RAPD-HRM with 3H Primer for E. Coli Screening.

A single random oligonucleotide 3H primer has been previously applied in random-amplified- polymorphic-DNA (RAPD)-PCR to distinguish stocked bacteria E. coli within a cocktail mixture also containing Enterococcus faecalis, Bifidobacterium longum and Ruminococcus gnavus. In this study, we demonstrate that a 702 base pair (bp) gene fragment can be amplified as a unique pattern by RAPD-PCR using a 3H primer in human faeces containing E. coli. This unique 702 bp amplicon contained a 687 bp gene fragment identified as the C-terminal region of the glutamate-ammonia-ligase adenyltransferase (glnE) gene of E. coli. By high-resolution melt (HRM) analysis, a mean melt-curve temperature of this 702 bp amplicon was determined to be approximately 88.1 ± 0.22 degrees Celsius (°C). A combination of RAPD with HRM in one single reaction based on this amplicon can achieve semi-quantitative detection of up to 102 CFU/ml of E. coli. To increase the signal intensity of HRM, a primer pair capable of screening E. coli directly from fresh human faeces was re-designed from the 687 bp gene segment, giving a mean peak melt-curve temperature at 88.35 ± 0.11 °C. Finally, single-nucleotide polymorphisms of this 687 bp gene segment were analysed for pathogenic E. coli strains, including UMN026, O83:H1, O104:H4, O157:H7 and O169:H41. We conclude that this 687 bp segment of the glnE gene has a high potential for screening of human faecal E. coli, including pathogenic strains, in contaminated food and water.

A novel framework to compare the effectiveness of ß-lactamase inhibitors against extended-spectrum ß-lactamase-producing Enterobacteriaceae.

OBJECTIVES: Extended-spectrum ß-lactamases (ESBLs) present a serious challenge in the treatment of Gram-negative bacterial infections. ESBLs mediate resistance to most ß-lactams, which may be reversed with the addition of an active ß-lactamase inhibitor (such as tazobactam, relebactam and avibactam). However, various ESBLs may exhibit different susceptibilities to these inhibitors, which could impact efficacy. We proposed a framework for comparing the efficacy of these inhibitors when combined with the same ß-lactam. METHODS: Three clinical isolates of Klebsiella pneumoniae harbouring CTX-M-15 and one Escherichia coli isolate with SHV-12 were examined. Piperacillin MICs were determined by broth dilution using escalating concentrations of tazobactam, relebactam and avibactam. The resulting MICs were characterized as response to inhibitor concentrations using an inhibitory sigmoid Emax model. Using the best-fit parameter values, the model was conditioned with fluctuating inhibitor concentrations to simulate instantaneous MICi profiles for each isolate-inhibitor pair. Using a simulated exposure of 4 g piperacillin every 8 h, %fT > MICi was estimated for each piperacillin/inhibitor combination. A hollowfibre infection model was subsequently used to validate the predicted effectiveness of selected combinations. RESULTS: In all scenarios, piperacillin MIC reductions were well characterized with increasing inhibitor concentrations. As predicted by %fT > MICi, combining piperacillin with avibactam (61.4%-73.6%) was found to be superior to tazobactam (13.5%-44.5%) for suppressing bacterial growth over time. CONCLUSION: We illustrated a practical approach to compare the performance of different inhibitors. This platform may be used clinically to identify the optimal pairing of various ß-lactams and ß-lactamase inhibitors for individual isolates producing ESBLs.

A rapidly fatal intracranial anaplastic hemangiopericytoma with de-novo dedifferentiation: emphasis on diagnostic recognition, molecular confirmation and discussion on treatment dilemma.

Solitary fibrous tumors/ hemangiopericytomas (SFT/HPC) are mesenchymal tumors that share a common genetic aberration and very rarely undergo dedifferentiation. We report a unique case of an intracranial anaplastic SFT/HPC with de-novo dedifferentiation, which pursued a rapidly fatal clinical course in a 41-year-old lady. The dedifferentiated component comprised a focal area of glandular formation with epithelial immunophenotype acquisition. The distinct biphasic pattern of the tumor imparted great diagnostic challenges to the pathologists. An increased awareness of SFT/HPCs with a diverse morphologic spectrum or even a biphasic histologic pattern is essential in working up such cases. We first attempted gamma knife radiosurgery in treating a recurrent dedifferentiated SFT/HPC; unfortunately it was to no avail. Although it is now known that SFT/HPC is characterized by NAB2-STAT6 gene fusion, the unavailability of targeted therapy against this molecular signature still results in a treatment dilemma.

Detection of Severe Acute Respiratory Syndrome-Like, Middle East Respiratory Syndrome-Like Bat Coronaviruses and Group H Rotavirus in Faeces of Korean Bats.

Bat species around the world have recently been recognized as major reservoirs of several zoonotic viruses, such as severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), Nipah virus and Hendra virus. In this study, consensus primer-based reverse transcriptase polymerase chain reactions (RT-PCRs) and high-throughput sequencing were performed to investigate viruses in bat faecal samples collected at 11 natural bat habitat sites from July to December 2015 in Korea. Diverse coronaviruses were first detected in Korean bat faeces, including alphacoronaviruses, SARS-CoV-like and MERS-CoV-like betacoronaviruses. In addition, we identified a novel bat rotavirus belonging to group H rotavirus which has only been described in human and pigs until now. Therefore, our results suggest the need for continuing surveillance and additional virological studies in domestic bat.

Inhibitory Effect of Imiquimod-Induced Psoriasis-Like Skin Inflammation in Mice by Histamine H4 Receptor Agonist 4-Methylhistamine.

Psoriasis is a chronic inflammatory immune-mediated autoimmune skin disorder. The histamine H4 receptor (H4R) agonist 4-methylhistamine (4-MH) plays an important role in immunomodulation of inflammatory responses associated with allergic inflammatory diseases. In this study, we investigated the effects of H4R agonist 4-MH on the development of imiquimod (IMQ)-induced psoriasis-like skin inflammation in mice and explored the immunoregulatory mechanism involved. The total clinical severity scores were significantly ameliorated by treatment with 4-MH (20 mg/kg) and 4-MH (40 mg/kg). Histological analysis of the skin revealed that 4-MH (20 mg/kg) and 4-MH (40 mg/kg) significantly attenuated the psoriatic phenotypes, including epidermal hyperplasis, hyperkeratosis and lymphocytes infiltration. Treatment with 4-MH (20 mg/kg) and 4-MH (40 mg/kg) led to reductions in the levels of Th1 cytokines (TNF-α, IFN-α, and IL-27) in the serum and dorsal skin, whereas Th17 cytokines levels (IL-17A and IL-23) did not change in response to treatment with 4-MH (20 mg/kg) and 4-MH (40 mg/kg). Furthermore, the number of CD4(+) CD25(+) FoxP3(+) regulatory T (Treg) cells was significantly increased by treatment with 4-MH (40 mg/kg). Taken together, these results imply that H4R agonist 4-MH might be an effective immunomodulatory approach for treatment of patients with psoriasis and the effects may be related to inhibited epidermal alteration, selectively reduced Th1 pro-inflammatory cytokines, and recruited CD4(+) CD25(+) FoxP3(+) Treg cells.

Peroxiredoxin II promotes hepatic tumorigenesis through cooperation with Ras/Forkhead box M1 signaling pathway.

The current study was carried out to define the involvement of Peroxiredoxin (Prx) II in progression of hepatocellular carcinoma (HCC) and the underlying molecular mechanism(s). Expression and function of Prx II in HCC was determined using H-ras(G12V)-transformed HCC cells (H-ras(G12V)-HCC cells) and the tumor livers from H-ras(G12V)-transgenic (Tg) mice and HCC patients. Prx II was upregulated in H-ras(G12V)-HCC cells and H-ras(G12V)-Tg mouse tumor livers, the expression pattern of which highly similar to that of forkhead Box M1 (FoxM1). Moreover, either knockdown of FoxM1 or site-directed mutagenesis of FoxM1-binding site of Prx II promoter significantly reduced Prx II levels in H-ras(G12V)-HCC cells, indicating FoxM1 as a direct transcription factor of Prx II in HCC. Interestingly, the null mutation of Prx II markedly decreased the number and size of tumors in H-ras(G12V)-Tg livers. Consistent with this, knockdown of Prx II in H-ras(G12V)-HCC cells reduced the expression of cyclin D1, cell proliferation, anchorage-independent growth and tumor formation in athymic nude mice, whereas overexpression of Prx II increased or aggravated the tumor phenotypes. Importantly, the expression of Prx II was correlated with that of FoxM1 in HCC patients. The activation of extracellular signal-related kinase (ERK) pathway and the expression of FoxM1 and cyclin D1 were highly dependent on Prx II in H-ras(G12V)-HCC cells and H-ras(G12V)-Tg livers. Prx II is FoxM1-dependently-expressed antioxidant in HCC and function as an enhancer of Ras(G12V) oncogenic potential in hepatic tumorigenesis through activation of ERK/FoxM1/cyclin D1 cascade.

Forced collapse of the blastocoel cavity improves developmental potential in cryopreserved bovine blastocysts by slow-rate freezing and vitrification.

This study was conducted to evaluate the effectiveness of forced collapse of the blastocoel before slow-rate freezing and vitrification of bovine blastocysts. Cryopreservation of bovine blastocysts has been proposed as a tool to improve the feasibility of cattle production using the embryo transfer technique. However, the low efficiency of frozen-thawed embryos survival and further development is a crucial problem. In this study, bovine in vitro and in vivo blastocysts were slow-rate frozen and vitrified after forced blastocoele collapse (FBC) of the blastocyst cavity by puncturing the blastocoele with a pulled Pasteur pipet. Differences in the developmental potential of frozen-thawed blastocysts derived from FBC and non-FBC groups were found in both slow-rate freezing and vitrification. Furthermore, we found that the total cell number of blastocysts in FBC groups was increased and the index of apoptosis in FBC groups was decreased. Consistent with these results, real-time RT-PCR analysis data showed that expression of the anti-apoptotic Bcl-XL gene was significantly increased by FBC groups, whereas expression of the pro-apoptotic Bax gene was significantly decreased by FBC groups. Our results also showed that pregnancy outcomes in both slow-rate frozen and vitrified bovine in vivo blastocysts could be improved by reducing the fluid content after FBC of the blastocyst cavity. Therefore, we suggest that FBC of the blastocyst cavity with a pulled Pasteur pipet is an effective pre-treatment technique for both slow-rate freezing and vitrification of bovine blastocysts.

Molecular cloning, characterization of porcine IZUMO1, an IgSF family member.

IZUMO1, belonging to the family of mammalian immunoglobulin proteins, has been well characterized in the mouse. Here, we describe the molecular cloning and expression analysis of porcine IZUMO1 (pIZUMO1). Partial sequence information published in the National Center for Biotechnology Information (NCBI) database was used to generate the full-length sequence for IZUMO1 using rapid amplification of cDNA ends (RACE). A search of the porcine genomic sequence in the NCBI database identified a bacterial artificial chromosome (BAC) encoding the pIZUMO1 gene. This BAC is derived from porcine chromosome 6 and is syntenic with the corresponding regions of mouse, bovine, and human genomes encoding the IZUMO gene family. This BAC was found to encode an IZUMO1 protein with a predicted amino acid sequence having high similarity with mouse and human IZUMO1. Western blot analysis of proteins from porcine tissues indicated that pIZUMO1 was specifically expressed in the sperm. Furthermore, to confirm whether pIZUMO1 forms complexes, we overexpressed pIZUMO1 in HEK293 cells. The recombinant pIZUMO1 from cell extracts was found to form complexes. Our finding suggests that pIZUMO1 forms homodimeric complex on the sperm membrane. Furthermore, an IVF inhibition assay with an antibody for the porcine IZUMO1 Ig-like domain showed that Ig-like domain effectively prevented pig sperm-egg interactions.

Tauroursodeoxycholic acid enhances the pre-implantation embryo development by reducing apoptosis in pigs.

Apoptosis is an important determinant of the normal development of pre-implantation embryos in vitro. Recently, endoplasmic reticulum (ER) stress-mediated apoptosis has been extensively investigated in a wide variety of diseases. Efficient functioning of the ER is essential for most cellular activities and survival. Tauroursodeoxycholic acid (TUDCA), an endogenous bile acid, has been reported to attenuate ER stress-mediated cell death by interrupting the classic pathways of apoptosis. Therefore, in this study, the anti-apoptotic effect of TUDCA on ER stress-induced apoptosis was examined in pre-implantation pig embryos. Also, tunicamycin was used to investigate the effects of ER stress on pig embryo development. After in vitro maturation and fertilization, presumptive pig embryos were cultured in NCSU-23 medium supplemented with TUDCA or TM for 6 days at 39 °C, 5% CO(2) in air. All data were analysed using one-way anova and Duncan's multiple range test in the statistical analysis system (SAS). In addition, we also determined the optimal TM and TUDCA concentrations. Samples were treated with TM at concentrations of 0, 1, 2 or 5 µm and with TUDCA at concentrations of 0, 100, 200 or 300 µm. When TM was used during in vitro culture, only 8.2% (8/97) of the embryos developed to the blastocyst stage when the treatment concentration was 1 µm compared with 27.4% (28/102) of the embryos in the control group (p < 0.05). In contrast, the frequency of blastocyst formation and the number of cells were higher when treated with 200 µm TUDCA compared with the control group (32.8% and 39.5 vs 22.2% and 35.6, p < 0.05). Moreover, the developmental rate to the blastocyst stage embryo in the group treated with TM and TUDCA was not significantly different from that of the control group (17.8%, 26/142 vs 24.9%, 36/145). Furthermore, the blastocyst cell number was enhanced (31.9 vs 36.9) and apoptosis reduced (TUNEL-positive nuclei number, 6.0 vs 3.2) by TUDCA treatment in pig embryos. In the real-time quantitative RT-PCR analysis, the expression of anti-apoptotic Bcl-XL gene was shown to be increased in the blastocyst stage because of TUDCA treatment, whereas expression of pro-apoptotic Bax was decreased. In addition, we also found that TUDCA decreased the rate of TM-induced apoptosis in the pre-implantation stage. Taken together, our results indicate that TUDCA improves the developmental competence of pig embryos by modulating ER stress-induced apoptosis during the pre-implantation stage.

Ependymal cyst in the cerebrum of an African green monkey (Chlorocebus aethiops).

A focal lesion was detected by magnetic resonance imaging in the right caudal occipital lobe of the cerebrum in an African green monkey (Chlorocebus aethiops). Neurological signs were not observed in this animal. At necropsy examination, an 8mm wedge-shaped intracranial cavity was found, which apparently did not communicate with the ventricles. Microscopically, the inner surface of the cavity was lined by ciliated cuboidal epithelium with positive immunoreactivity for S100 protein, glial fibrillary acidic protein and cytokeratin. Based on the gross, microscopical and immunohistochemical findings the lesion was classified as an ependymal cyst. To the best of our knowledge, this is the first report of an ependymal cyst in an African green monkey.

Pathological examination of the placenta: raison d'être, clinical relevance and medicolegal utility.

Formal pathological examination of the placenta provides valuable information to the obstetrician, neonatologist, paediatrician and family. This article aims to provide the clinician with an overview of the significance of placental examination in relation to common or important pathological processes, and the utility of information obtained therein in explaining adverse outcomes, management of subsequent pregnancies, and assessment of newborn risk for the development of short- or long-term sequelae. General guidelines for placental examination, and logistical and practical issues are also discussed. Finally, the role of the placenta in the defence of obstetricians and other healthcare workers in cases of poor neonatal outcome is described.

The late-stage foetal liver microenvironment is essential for later sensitivity of B-lymphopoiesis to suppression by oestrogens.

B-lymphopoiesis in FL differs notably from that of adult B-lymphopoiesis in being resistant to suppression by oestrogens due to the lack of expression of oestrogen receptors in B-cell progenitors and precursors. We have transplanted middle-stage FL cells (E14.5) to adult male mice and demonstrated that B-lymphopoiesis derived from FL cells remained resistant to suppression by exogenous oestrogen for several months compared to adult BM cells. This significant difference strongly suggests that the latestage FL environment exerts an inductive action on the haematopoietic stem cells and is mandatory for later sensitivity of B-lymphopoiesis to suppression by oestrogens. The results also provide the first in vivo functional confirmation of a differential responsiveness of FL- and adult BM-derived B-lymphopoiesis to suppression by oestrogens.

Comparison of the optical coherence tomographic features of choroidal neovascular membranes in pathological myopia versus age-related macular degeneration, using quantitative subanalysis.

AIM: To compare the retinal morphological characteristics of eyes with choroidal neovascularisation (CNV) secondary to pathological myopia versus eyes with CNV secondary to age-related macular degeneration (AMD), using quantitative optical coherence tomography (OCT) subanalysis. METHODS: Twenty-one eyes of 21 patients newly diagnosed as having CNV secondary to pathological myopia, and 43 consecutive cases of eyes with newly diagnosed subfoveal CNV secondary to AMD were retrospectively collected. In all patients, StratusOCT images and fluorescein angiograms (FA) were available for analysis. StratusOCT images were analysed using custom software (termed "OCTOR"), which allowed calculation of the thickness/volume of the neurosensory retina, subretinal fluid (SRF), subretinal tissue (SRT) and pigment epithelial detachments (PEDs). FA images were used to calculate CNV leakage area and CNV lesion size for each eye. RESULTS: The total volume of neurosensory retina in the pathological myopia group was significantly less than in the AMD group (7.10 (SD 0.50) mm3 vs 7.76 (0.93) mm3, p = 0.004). The total volume of SRF in the pathological myopia group was less than in the AMD group, but the difference was not statistically significant (0.33 (1.38) mm3 vs 0.55 (0.82) mm3, p = 0.434). The total volume of SRT in the pathological myopia group was less than in the AMD group, but the difference was not statistically significant (0.16 (0.15) mm3 vs 0.36 (0.60) mm3, p = 0.144). The total volume of PED in the pathological myopia group was markedly less than in the AMD group (0.01 (0.03) mm3 vs 1.09 (1.89) mm3, p<0.001). On FA, the total leakage of CNV in the AMD group was significantly greater than in the pathological myopia group (4.17 (3.29) DAs vs 0.53 (0.58) DAs, p<0.001). CONCLUSIONS: CNV lesions in pathological myopia were associated with considerably less retinal oedema, SRF and SRT compared with CNV associated with AMD. PEDs were almost negligible in myopic lesions compared with AMD. These findings are consistent with previous clinical and angiographic descriptions of myopic CNV as relatively small lesions with modest exudation.

Transcriptional regulation of GSDML gene by antisense-oriented HERV-H LTR element.

During the course of hominoid evolution, a new transcript variant of the GSDML (gasdermin-like protein) gene was formed by the integration of the antisense-oriented HERV-H (human endogenous retrovirus) LTR (long terminal repeat) element. To investigate regions that are critical for transcriptional regulation of the GSDML gene, we generated seven deletion mutants from a full-length clone (clone 1/630) that includes the HERV-H LTR sequence and compared their expression levels relative to the full-length parental clone using a transient transfection assay. In the transient transfection assay, deletion of the 5' flanking region (cellular origin) of the HERV-H LTR sequence led to a 4.5-fold increase in expression compared to the full-length clone, while deletion of the U5 region showed a significant decrease in transcriptional activity. Deletion of the 3' flanking region of the LTR sequence (clone 42/451) showed similar transcriptional activity to a clone missing the 5' flanking region of cellular origin (clone 42/630). Taken together, these data indicate that the HERV-H LTR sequence (viral origin) positively regulates transcriptional activity of the GSDML gene and that the 5' flanking region sequence (cellular origin) exerts negative transcriptional regulation.

Pharmacokinetics and tissue distribution of 3-((5-(6-methylpyridin-2-yl)-4-(quinoxalin-6-yl)-1H-imidazol-2-yl)methyl)benzamide; a novel ALK5 inhibitor and a potential anti-fibrosis drug.

The authors investigated the pharmacokinetics and metabolism of 3-((5-(6-methylpyridin-2-yl)-4-(quinoxalin-6-yl)-1H-imidazol-2-yl)methyl)benzamide (IN-1130), a novel ALK5 inhibitor, which suppresses renal and hepatic fibrosis, and also exerts anti-metastatic effects on breast cancer-bearing MMTV-cNeu mice model. Plasma half-lives of orally administered IN-1130 were 62.6 min in mice, 76.6 +/- 10.6 min in dogs, 156.1 +/- 19.3 min in rats, and 159.9 +/- 59.9 min in monkeys. IN-1130 showed a high apparent permeability coefficient (P(app)) of (45.0 +/- 2.3) x 10(-6) cm s(-1) in in vitro permeability tests in a Caco-2 cell monolayer model. The bioavailability of orally administered IN-1130 was 84.9% in dogs and 34.4% in monkeys (oral dose, 5.5 mg kg(-1)), 11.4% in rats and 8.95% in mice (oral dose, 50.3 mg kg(-1)), respectively. Orally given IN-1130 was readily distributed into liver, kidneys and lungs. The major metabolite of IN-1130 (M1) was detected in the systemic circulation of rat and mouse and was purified and tentatively identified as 3-((4-(3-hydroxyquinoxaline-6-yl)-5-(6-methylpyridine-2-yl)-1H-imidazol-2-yl)methyl)benzamide or 3-((4-(2-hydroxyquinoxalin-6-yl)-5-(6-methylpyridine-2-yl)-1H-imidazol-2-yl)methyl)benzamide. The highest levels of M1 were found in liver. The results of this study suggest that IN-1130 has the potential to serve as an effective oral anti-fibrotic drug.

Intraductal papillary mucinous tumour of the pancreas: differentiation of malignancy and benignancy by CT.

AIM: To retrospectively identify signs predictive of malignant intraductal papillary mucinous tumour (IPMT) of the pancreas on computed tomography (CT) images. MATERIALS AND METHODS: Thirty-four benign and 21 malignant pancreatic IPMTs were evaluated. Preoperative helical CT images in these 55 cases of pathologically proven pancreatic IPMT were reviewed by two radiologists unaware of the histological grading. Tumour morphological types, locations, numbers and sizes of cystic lesions, maximum main pancreatic duct diameters, the presence of septa, mural nodule, wall thickening, and calcification in cysts, communication with the main pancreatic duct, peripancreatic haziness, protrusion of duodenal papilla, pancreatic atrophy, lymphadenopathy and distant metastasis were analysed using univariate and multivariate analysis. RESULTS: Main duct IPMTs were more likely to be malignant (71%) than branch duct (23%) or combined type IPMTs (28%; p=0.002). Among the branch duct type and combined types, large cystic lesion (p=0.018), the presence of a mural nodule (p=0.018), a thickened wall (p=0.009), and peripancreatic haziness (p=0.039) were found to predict malignancy. CONCLUSION: CT is helpful in the preoperative differentiation of malignant and benign pancreatic IPMT. The presence of a dilated main pancreatic duct, mural nodules, thickened wall and peripancreatic haziness may be used as independent predictive signs of malignancy.

Cytokine gene expression in regenerating haematopoietic tissues of mice after cyclophosphamide treatment.

The goal of the study was to investigate changes in expression of selected growth factors tentatively involved in regeneration of haematopoietic tissues (bone marrow and spleen) following cyclophosphamide (CY) damage in the mouse. The bone marrow (BM) and spleen were examined separately, since the regenerating pattern for haematopoietic progenitor cells (HPC) markedly differs in these two haematopoietically active organs after CY. Cytokines assumed to have a stimulatory effect on HPC - stem cell factor (SCF), fetal liver tyrosine kinase 3-ligand (flt3-ligand), thrombopoietin (TPO), stromal cell-derived factor 1 (SDF-1), oncostatin M (OSM) -, a suppressive effect on HPC proliferation - macrophage inflammatory protein-1alpha (MIP-1alpha), transforming growth factor-beta1 (TGFbeta1), tumour necrosis factor-alpha (TNFalpha) - and to be involved in migration of HPC (SCF, flt3-ligand, MIP1alpha, SDF-1) were examined at the level of mRNA expression by means of real-time RT-PCR. The expression of a particular cytokine appears to be similar in both BM and spleen of untreated mice. CY administration changed the expression pattern of the studied genes in BM and spleen. In BM, the levels of mRNAs for SCF and SDF-1 were increased and that for TGFbeta1 decreased at time intervals at which HPC are known to proliferate intensively during BM regeneration. In contrast, stimulated proliferation of HPC in spleen was accompanied by increased expression of flt3-ligand and oncostatin M. Upon mobilization of HPC from BM into blood after CY, the expression of SCF, TPO, SDF-1 and TGFbeta1 tends to decrease in BM. Accumulation of HPC in spleen is accompanied by increased mRNA for flt3-ligand and OSM. Our findings demonstrate that different cytokines may be involved in the proliferation and mobilization/homing of HPC during recovery after CY damage in BM and spleen.