AAV8 vector expressing IL24 efficiently suppresses tumor growth mediated by specific mechanisms in MLL/AF4-positive ALL model mice

Mixed-lineage leukemia (MLL)/AF4-positive acute lymphoblastic leukemia (ALL) is a common type of leukemia in infants, which is associated with a high relapse rate and poor prognosis. IL24 selectively induces apoptosis in cancer cells and exerts immunomodulatory and antiangiogenic effects. We examined the effects of adeno-associated virus type 8 (AAV8) vector-mediated muscle-directed systemic gene therapy in MLL/AF4-positive ALL using IL24. In a series of in vitro studies, we examined the effects of AAV8-IL24-transduced C2C12 cell-conditioned medium. We also examined the effects of AAV8-IL24 in MLL/AF4 transgenic mice. The results revealed the effects of AAV8-IL24 in MLL/AF4-positive ALL both in vitro and in vivo. With regard to the mechanism of therapy using AAV8-IL24 in MLL/AF4-positive ALL, we demonstrated the antiangiogenicity and effects on the ER stress pathway and unreported pathways through inhibition of S100A6 and HOXA9, which is specific to MLL/AF4-positive ALL. Inhibition of S100A6 by IL24 was dependent on TNF-α and induced acetylation of p53 followed by activation of the caspase 8-caspase 3 apoptotic pathway. Inhibition of HOXA9 by IL24, which was independent of TNF-α, induced MEIS1 activation followed by activation of the caspase 8-caspase 3 apoptotic pathway. Thus, gene therapy using AAV8-IL24 is a promising treatment for MLL/AF4-positive ALL.     

A case of blastic plasmacytoid dendritic cell neoplasm: Cytomorphological findings of the touch imprint specimen of lymph node

The patient is a 75‐year‐old man with axillary lymphadenopathy presenting an indurated papule on his buttock. Touch imprint cytology of the biopsied axillary lymph node revealed the monotonous appearance of medium‐sized tumor cells. The nuclei had a slightly irregular contour, finely dispersed chromatin, and a conspicuous nucleolus. Some tumor cells had intracytoplasmic microvacuoles. Immunohistochemistry of the imprint specimens showed that the tumor cells were positive for CD56 and CD123. Histological diagnosis of the lesion was blastic plasmacytoid dendritic cell neoplasm (BPDCN). Epstein‐Barr virus‐encoded RNAs were not detected in the tumor cells. Neither immunoglobulin heavy chain genes nor T‐ cell receptor genes was clonally rearranged. BPDCN should be strongly considered during the differential diagnosis of CD56‐positive neoplasms of the skin. We demonstrated a possible contribution of the cytomorphological and immunohistochemical findings of the touch imprint specimens to the diagnosis of BPDCN. Diagn. Cytopathol. 2013. © 2012 Wiley Periodicals, Inc.     

Use of a triple antibiotic solution affects the healing process of intentionally delayed replanted teeth in mice

ObjectiveA mixture of ciprofloxacin, metronidazole, and minocycline (3Mix) has been reported to be effective against oral bacteria from carious and endodontic lesions in vitro and in vivo. The objective of this study was to establish an animal model using mice for the application of 3Mix following intentionally delayed tooth replantation and to investigate the effects of 3Mix on the healing process of dental pulp and periodontal tissues.MethodsUpper first molars of ICR mice were extracted, immersed in 3Mix solution at different concentrations for 5–60 min with or without the use of a transfer solution (phosphate buffer solution (PBS)), in addition to transfer solution alone, and subsequently repositioned in the sockets. Immunohistochemistry for nestin and Ki-67, histochemistry for TRAP, and TUNEL assay were performed to assess pulpal healing during days 7–21.ResultsIncreased apoptosis was observed in the PBS group at week 1, followed by cell proliferation at week 2, and tertiary dentin and/or bone-like tissue formation at week 3. In contrast, nestin-positive, newly differentiated, odontoblast-like cells began to align along the pulp–dentin border following the appearance of Ki-67- and TUNEL-positive cells during weeks 1–2 in the 3Mix groups, suggesting that pulpal healing was accelerated. Severe root ankylosis was observed exclusively in the 3Mix groups. Rinsing with PBS before replantation partially rescued the viability of the periodontal ligament, but pulpal healing was delayed.ConclusionsThe application of 3Mix promotes pulpal regeneration of intentionally delayed replanted teeth; however, its use may induce severe damage to periodontal tissues.     

Impaired regulatory T cell reconstitution in patients with acute graft-versus-host disease and cytomegalovirus infection after allogeneic bone marrow transplantation

To elucidate the correlation between regulatory T cells (Tregs) and acute graft-versus-host disease (aGVHD) or cytomegalovirus infection following allogeneic bone marrow transplantation (allo-BMT), we evaluated either CD4?CD25(high) or FOXP3? Treg-enriched cells in peripheral blood (PB) from 20 patients who received allo-BMT, and in biopsies of skin with aGVHD. Proportions of CD4?CD25(high)FOXP3? cells in total lymphocytes, but not other types of T cells, were lower in patients who eventually developed grades II-IV aGVHD (n = 13) than in others (n = 7, P < 0.001). Proportions of CD62L? cells in CD4?CD25(high) cells at day +30 were lower (P < 0.01) in patients who eventually showed cytomegalovirus viremia (n = 6) than in others (n = 14). Incidence of aGVHD (P < 0.05) or cytomegalovirus viremia (P < 0.05) was higher in patients without these complications, but with lower proportions of PB CD4?CD25(high)FOXP3? cells at day +30 (n = 8) than in others (n = 8). However, in skin with aGVHD (n = 5), there was marked or slightly increased infiltration of CD8? cells (P < 0.001) or CD3?FOXP3? cells (P < 0.05), respectively, when compared with control (n = 5), resulting in threefold higher ratio of CD8?/CD3?FOXP3? cells in aGVHD relative to controls (P < 0.05). Thus, impaired reconstitution of Tregs may be associated with aGVHD and CMV infection. Moreover, imbalance of Tregs and CD8? cells may play a role in aGVHD tissue.     

Effectiveness and safety of adalimumab in Japanese patients with rheumatoid arthritis: retrospective analyses of data collected during the first year of adalimumab treatment in routine clinical practice (HARMONY study)

We retrospectively investigated the ability of adalimumab (ADA) to reduce disease activity, improve physical function, and retard the progression of structural damage in 167 patients with rheumatoid arthritis. Clinical and functional outcomes were compared between patients with or without prior biologic treatment and those with or without concomitant methotrexate (MTX) treatment. At week 52, 38.3% achieved clinical remission: 42.4 and 28.6% of patients achieved remission in those without and with previous biologics, respectively, while 42.7 and 12.5% of patients achieved remission in those with and without concomitant MTX, respectively. ADA treatment significantly reduced the rate of radiographic progression from 27.1 ± 46.0 (median 13.6; 25th-75th percentiles 8.3 to 28.9) at baseline to 0.8 ± 5.0 (median 0.0; 25th-75th percentiles -0.9 to 2.0) at week 52 (P < 0.0001). Radiographic progression was absent in 59.8% of patients. Sixty adverse events (34.21/100 patient-years) were reported, 16 of which were serious (9.12/100 patient-years). ADA therapy is highly effective for reducing disease activity, improving physical function, and limiting radiographic progression. It is generally safe and well tolerated by Japanese RA patients in routine clinical practice.     

TGF-β1 Promotes Lymphangiogenesis during Peritoneal Fibrosis

Peritoneal fibrosis (PF) causes ultrafiltration failure (UFF) and is a complicating factor in long-term peritoneal dialysis. Lymphatic reabsorption also may contribute to UFF, but little is known about lymphangiogenesis in patients with UFF and peritonitis. We studied the role of the lymphangiogenesis mediator vascular endothelial growth factor-C (VEGF-C) in human dialysate effluents, peritoneal tissues, and peritoneal mesothelial cells (HPMCs). Dialysate VEGF-C concentration correlated positively with the dialysate-to-plasma ratio of creatinine (D/P Cr) and the dialysate TGF-β1 concentration. Peritoneal tissue from patients with UFF expressed higher levels of VEGF-C, lymphatic endothelial hyaluronan receptor-1 (LYVE-1), and podoplanin mRNA and contained more lymphatic vessels than tissue from patients without UFF. Furthermore, mesothelial cell and macrophage expression of VEGF-C increased in the peritoneal membranes of patients with UFF and peritonitis. In cultured mesothelial cells, TGF-β1 upregulated the expression of VEGF-C mRNA and protein, and this upregulation was suppressed by a TGF-β type I receptor (TGFβR-I) inhibitor. TGF-β1–induced upregulation of VEGF-C mRNA expression in cultured HPMCs correlated with the D/P Cr of the patient from whom the HPMCs were derived (P<0.001). Moreover, treatment with a TGFβR-I inhibitor suppressed the enhanced lymphangiogenesis and VEGF-C expression associated with fibrosis in a rat model of PF. These results suggest that lymphangiogenesis associates with fibrosis through the TGF-β–VEGF-C pathway.The decrease in ultrafiltration capacity that is associated with the high peritoneal solute transport that is observed after prolonged peritoneal dialysis (PD) treatment is a major reason for its discontinuation.^1–4 Several studies have shown that a higher peritoneal solute transport rate is associated with reduced survival of PD patients.^1,2,5 The characteristic features of chronic peritoneal damage in PD treatment are associated with submesothelial fibrosis and neoangiogenesis.^6,7 Analyses of the surface peritoneum showed no significant changes in vessel density with duration of PD.^6,8 In addition, the vessel density in patients with ultrafiltration failure (UFF) was significantly higher than the vessel density in normal individuals or non-PD patients, but it was not higher than the vessel density in patients undergoing PD.⁶ These findings suggest that factors other than increased vascular density may be involved in disease states associated with increased transport of peritoneal membranes. In addition, the relationship between peritoneal fibrosis and UFF remains obscure.Blood capillaries have a continuous basal lamina with tight interendothelial junctions and are supported by pericytes and smooth muscle cells. In contrast, lymphatic capillaries are thin-walled with a wide lumen and do not contain pericytes or basement membrane. The structures of lymphatic vessels are suitable for the removal of tissue fluid, cells, and macromolecules from the interstitium.^9–11 If lymphangiogenesis develops in the peritoneal membrane, absorption of the PD fluid could be increased and lead to UFF. An increase in the number of lymphatic vessels has recently been reported in several disease conditions, including tumor metastasis,^12–15 chronic respiratory inflammatory diseases,^16–18 wound healing,¹⁹ and renal transplant rejection.^20,21 We recently reported that lymphangiogenesis had developed in tubulointerstitial fibrosis of human renal biopsy specimens,²² and we also reported the mechanisms of lymphangiogenesis in rat unilateral ureteral obstruction models.²³The lymphatic absorption rate, which is measured by the rate at which intraperitoneally administered radioactive serum albumin or macromolecule dextran 70 disappears, is significantly higher in patients with UFF, and lymphatic reabsorption is considered to be one of the causes of UFF.^24–27 However, the results from these clinical approaches have been controversial.^28,29 In addition, little is known about the pathology and the process of lymphangiogenesis in patients with UFF and peritonitis.In this study, we investigated lymphangiogenesis and the expression of vascular endothelial growth factor-C (VEGF-C), which is a potentially important mediator of lymphangiogenesis, in human peritoneal tissues, PD effluent, and peritoneal mesothelial cells. We also explored VEGF-C induction by TGF-β1 in the human mesothelial cell line (Met-5A) and cultured human peritoneal mesothelial cells (HPMCs) from the spent PD effluent of patients with varying rates of peritoneal transport. Finally, we explored the relationship between peritoneal fibrosis and lymphangiogenesis in rats that were administered chlorhexidine gluconate (CG) into the abdominal cavity, which provides a model of chemically induced peritoneal inflammation/fibrosis.^30–32 This work is the first report to show that lymphangiogenesis is linked to the peritoneal fibrosis that is often associated with a high peritoneal transport rate.     

Perineural Invasion in Extrahepatic Cholangiocarcinoma: Prognostic Impact and Treatment Strategies

Background

The significance of perineural invasion in extrahepatic cholangiocarcinoma has not been fully elucidated. This study aims to determine the prognostic impact of and optimal treatment strategy for perineural invasion in patients with extrahepatic cholangiocarcinoma.

Methods

Medical records of 133 patients with extrahepatic cholangiocarcinoma who underwent curative resection were reviewed retrospectively. Ninety-eight patients had perineural invasion and 35 patients did not. Univariate and multivariate survival analyses were performed to clarify the prognostic impact of and optimal treatment strategy for perineural invasion.

Results

Only tumor differentiation (P?=?0.024) was independently associated with perineural invasion in the multivariate logistic regression model. Multivariate survival analysis revealed that perineural invasion (P?=?0.002), resection margin status (P?=?0.016), and International Union Against Cancer (UICC) pT factor (P?=?0.015) were independent prognostic factors of overall survival. Overall 5-year survival rates for patients with and without perineural invasion were 28 and 74 %, respectively. Among 98 patients with perineural invasion, the use of adjuvant chemotherapy (P?=?0.003), lymph node status (P?=?0.015), resection margin status (P?=?0.008), and UICC pT factor (P?=?0.016) were independently associated with overall survival by multivariate analysis. Overall 5-year survival rates for patients with perineural invasion who did and did not receive adjuvant chemotherapy were 33 and 21 %, respectively (P?=?0.023).

Conclusions

Perineural invasion is a potent prognostic factor in extrahepatic cholangiocarcinoma. Adjuvant chemotherapy may improve the overall survival of patients with perineural invasion.     

Effects of eldecalcitol on bone and skeletal muscles in glucocorticoid-treated rats

Glucocorticoids cause secondary osteoporosis and myopathy, characterized by type II muscle fiber atrophy. We examined whether a new vitamin D₃ analogue, eldecalcitol, could inhibit glucocorticoid-induced osteopenia or myopathy in rats, and also determined the effects of prednisolone (PSL) and/or eldecalcitol on muscle-related gene expression. Six-month-old female Wistar rats were randomized into four groups: PSL group (10 mg/kg PSL); E group (0.05 µg/kg eldecalcitol); PSL + E group; and control group. PSL, eldecalcitol, and vehicles were administered daily for 2 or 4 weeks. Right calf muscle strength, muscle fatigue, cross-sectional areas (CSAs) of left tibialis anterior muscle fibers, and bone mineral density (BMD) were measured following administration. Pax7, MyoD, and myogenin mRNA levels in gastrocnemius muscles were also determined. Muscle strength was significantly higher in the PSL + E group than in the PSL group (p < 0.05) after 4 weeks, but not after 2 weeks. No significant difference in muscle fatigue was seen between groups at 2 or 4 weeks. CSAs of type II muscle fibers were significantly larger in the E group and the PSL + E group than in the PSL group at 4 weeks (p = 0.0093, p = 0.0443, respectively). Eldecalcitol treatment for 4 weeks maintained the same BMD as the PSL + E group. After 2 weeks, but not 4 weeks, eldecalcitol treatment significantly increased Pax7 and myogenin mRNA expression in gastrocnemius muscle, and PSL also stimulated myogenin expression. Eldecalcitol appears to increase muscle volume and to protect against femur BMD loss in PSL-administered rats, and it may also stimulate myoblast differentiation into early myotubes.     

The feasibility of laparoscopic extended pelvic surgery for rectal cancer

Purpose

The present study aimed to assess the safety and feasibility of laparoscopic extended pelvic surgery for primary or recurrent rectal cancer.

Methods

The data on 77 patients, who underwent extended pelvic surgery between February 2008 and June 2014, were retrospectively analyzed. The patients were divided, based on their treatment history, into an open surgery (OS) group (n = 41) and a laparoscopic surgery (LS) group (n = 36).

Results

The operative time in the LS group was significantly longer than that in the OS group (766 vs. 561 min; p < 0.001). In contrast, the LS group was associated with a significantly lower volume of intraoperative blood loss (195 vs. 923 ml; p < 0.001), fluid balance (5.38 vs. 8.23 ml/kg/h; p < 0.001) and rate of complications (40.0 vs. 68.3 %; p = 0.035), and a significantly shorter postoperative hospital stay. The postoperative levels of colloid osmotic pressure and albumin were significantly higher in the LS group.

Conclusion

The operative time of the LS group was longer than that of the OS group; however, the LS group experienced less blood loss and fewer complications. Moreover, LS was associated with a reduction in intraoperative infusions and a reduced fluid balance, which maintained homeostasis.