Suppression of STAT5b in pancreatic cancer cells leads to attenuated gemcitabine chemoresistance, adhesion and invasion.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal solid malignancies, and there is an urgent need for new therapeutic strategies based on the molecular biology of PDAC. Signal transducers and activators of transcription 5 (STAT5) are known to be activated in a variety of malignancies and involved in tumor proliferation, apoptosis, and invasion, whereas the expression and biological role of STAT5b in PDAC are less clearly defined. In the present study, we examined the expression and role of STAT5b in human pancreatic cancer cell lines. Expressions of STAT5b mRNA and protein were detected in eight kinds of pancreatic cancer cells. Confocal microscopy and western blot analysis indicated that STAT5b is localized in both cytoplasm and nuclei. Immunoprecipitation analysis revealed tyrosine phosphorylation of STAT5b in pancreatic cancer cells. These results indicate that STAT5b in pancreatic cancer cells is constitutively activated. STAT5b shRNA clones in PANC-1 cells, which express relatively high levels of STAT5b, exhibited reduced chemoresistance against gemcitabine, adhesion and invasion compared to sham. On the other hand, AsPC-1 and BxPC3 cells, which express relatively low levels of STAT5b, exhibited reduced chemoresistance compared to PANC-1 cells. Moreover, STAT5b overexpression clones in AsPC-1 cells exhibited increased chemoresistance compared to sham. STAT5b shRNA clones in PANC-1 cells were more sensitive to the proapoptotic actions of gemcitabine, as evidenced by PARP and cleaved caspase-3 activation. Gemcitabine also significantly reduced Bcl-xL levels in the STAT5b shRNA-expressing cells. We also investigated the clinicopathological characteristics of STAT5b expression of PDAC. Although a significant correlation between STAT5b expression and overall survival rates was not observed, a significant correlation with main pancreatic duct invasion was observed. These findings suggest that STAT5b confers gemcitabine chemoresistance and promotes cell adherence and invasiveness in pancreatic cancer cells. Targeting STAT5b may lead to novel therapeutic strategies for PDAC.

Clinical outcomes for 14 consecutive patients with solid pseudopapillary neoplasms who underwent laparoscopic distal pancreatectomy.

INTRODUCTION: The postoperative results of laparoscopic distal pancreatectomy for solid pseudopapillary neoplasm of the pancreas (SPN), including the effects of spleen-preserving resection, are still to be elucidated. METHODS: Of the 139 patients who underwent laparoscopic pancreatectomy for non-cancerous tumors, 14 consecutive patients (average age, 29.6 years; 1 man, 13 women) with solitary SPN who underwent laparoscopic distal pancreatectomy between March 2004 and June 2015 were enrolled. The tumors had a mean diameter of 4.8 cm. Laparoscopic spleen-preserving distal pancreatectomy was performed in eight patients (spleen-preserving group), including two cases involving pancreatic tail preservation, and laparoscopic spleno-distal pancreatectomy was performed in six patients (standard resection group). RESULTS: The median operating time was 317 min, and the median blood loss was 50 mL. Postoperatively, grade B pancreatic fistulas appeared in two patients (14.3%) but resolved with conservative treatment. No patients had postoperative complications, other than pancreatic fistulas, or required reoperation. The median postoperative hospital stay was 11 days, and the postoperative mortality was zero.None of the patients had positive surgical margins or lymph nodes with metastasis. The median follow-up period did not significantly differ between the two groups (20 vs 39 months, P = 0.1368). All of the patients are alive and free from recurrent tumors without major late-phase complications. CONCLUSION: Laparoscopic distal pancreatectomy might be a suitable treatment for patients with SPN. A spleen-preserving operation is preferable for younger patients with SPN, and this study demonstrated the non-inferiority of the procedure compared to spleno-distal pancreatectomy.

Mitotic index and multipolar mitosis in routine histologic sections as prognostic markers of pancreatic cancers: A clinicopathological study.

OBJECTIVES: Pancreatic cancer is characterized by genomic complexity and chromosomal instability, and atypical mitotic figures are morphological features of this phenotype. In the present study, we determined the frequency and the clinicopathological and prognostic significance of mitotic figures in pancreatic cancers. METHODS: We surveyed the mitotic figures of the normal ductal epithelium, acinar cells, pancreatic intraepithelial neoplasias, and pancreatic cancers on hematoxylin-and-eosin-stained tissue specimens (n = 121). RESULTS: Pancreatic cancer cells showed significantly higher mitotic indices as compared with the ductal cells, acinar cells, and pancreatic intraepithelial neoplasias. Both normal and atypical mitosis were significantly elevated only in pancreatic cancers. In pancreatic cancers, approximately 30% of total mitosis was atypical including multipolar, lag-type, ring and asymmetrical mitosis, and anaphase bridges. The Kaplan-Meier curves in pancreatic cancers showed significant correlations between total mitosis and disease free survival. Furthermore, the cases with multipolar mitosis showed poorer prognosis than those without. Lymph node metastasis and multipolar mitosis were independent prognostic factors for overall survival of patients with pancreatic cancer. In addition, lymph node metastasis and total mitosis were independent factors for disease free survival. CONCLUSION: These findings suggest that routinely obtained pathological specimens, even small biopsy or cytological specimens, can provide valuable information concerning the prognosis of pancreatic cancers.

Clinical outcomes of 15 consecutive patients who underwent laparoscopic insulinoma resection: The usefulness of monitoring intraoperative blood insulin during laparoscopic pancreatectomy.

BACKGROUND: Insulinoma is a very serious functional tumor. Surgeons should confirm complete resection of insulinomas before completing the operation, even in laparoscopic surgery. METHODS: Between August 2007 and September 2014, 15 consecutive patients with biochemical evidence of an insulinoma underwent laparoscopic pancreatectomy. Intraoperatively, a peripheral arterial blood sample was taken, and insulin was measured by quick insulin assay. Insulin levels were determined before anesthesia induction, every 30 min thereafter, and every 30 min for at least 1 h after tumor resection to confirm insulin levels did not increase before surgery was completed. RESULTS: All 15 patients (3 men and 12 women, average age 57.2 years) successfully underwent laparoscopic resection. One patient had two tumors, and the remaining 14 patients had one tumor each (three in the head, five in the body, and eight in the tail of the pancreas). Preoperative localization and regionalization studies identified the tumor correctly through CT (12/15 [80.0%]), MRI (9/12 [75.0%]), angiography (11/13 [84.6%]), endoscopic ultrasonography (7/10 [70.0%]), and selective arterial calcium injection (14/14 [100%]). Intraoperative ultrasonography detected 13 of 15 tumors (86.7%), and intraoperative blood insulin monitoring confirmed the complete resection of 16 of 16 tumors (100%). All patients were discharged with normal insulin levels and have been followed up for 3-88 months. There has been no recurrence of symptoms in any patients and none has died. CONCLUSION: Complete removal of an insulinoma can be reliably predicted by intraoperative blood insulin monitoring even in laparoscopic pancreatectomies.

Laparoscopic distal pancreatectomy: Educating surgeons about advanced laparoscopic surgery.

INTRODUCTION: Laparoscopic distal pancreatectomy (Lap-DP) has been recognized worldwide as a feasible and highly beneficial procedure. The aim of this study is to investigate whether Lap-DP techniques are being implemented safely by surgeons training to perform this procedure. METHODS: We retrospectively compared the perioperative outcomes of Lap-DP in patients operated on by the surgeon originating this procedure at our hospital (expert surgeon group [E group], n = 47) and patients operated on by surgeons training to perform this procedure (training surgeons group [T group], n = 53). RESULTS: The median operating times for the E group and T group were 321 min (range, 150-653 min) and 314 min (range, 173-629 min), respectively; these times were not significantly different (P = 0.4769). The median blood loss in the T group (100 mL; range, 0-1950 mL) was significantly smaller than in the E group (280 mL; range, 0-1920 mL) (P = 0.0003). There were no significant intergroup differences in other operative results: combined operation ratio, spleen- and splenic vessels-preserving ratio, hand-assisted procedure ratio, and the ratio of transition to open. The frequency of pancreatic fistulas in the E group and T group was 12.8% and 16.9%, respectively; these rates were not significantly different (P = 0.5886). There were no significant differences between the two groups in terms of other complications and reoperation rates. The median hospital stay for the E group was significantly shorter than for the T group (10 vs 13 days; P = 0.0307). CONCLUSION: This retrospective analysis shows that teaching safe Lap-DP techniques to surgeons is reflected in stable perioperative outcomes.

[Radical resection of a locally advanced pancreatic tail adenosquamous carcinoma treated with S-1 and gemcitabine as neoadjuvant chemotherapy - a case report].

We report a case of locally advanced pancreatic tail adenosquamous carcinoma that was treated by performing R0 resection after neoadjuvant chemotherapy with S-1 and gemcitabine. A 75-year-old man visited our hospital because of left lateral abdominal pain. On the basis of computed tomography and endoscopic biopsy findings, an 80-mm locally advanced pancreatic tail carcinoma with direct invasion to the gastric upper body, splenic flexure of the colon, and left kidney was diagnosed. Combined chemotherapy with S-1 and gemcitabine was initiated for reduction in the tumor size. After 11 courses of treatment, computed tomography revealed a partial response in tumor size reduction. Grade 3 neutropenia was observed as an adverse event. Distal pancreatectomy, proximal gastrectomy, partial resection of the descending colon, resection of the left kidney and left adrenal gland, and D2 lymph node dissection were performed. The pathological diagnosis was adenosquamous carcinoma in the pancreatic tail, and an R0 resection was achieved. However, a month after surgery, multiple distant liver metastases were observed. Neoadjuvant chemotherapy with S-1 and gemcitabine may reduce the tumor size in locally advanced pancreatic tail adenosquamous carcinoma and increase the R0 resection rate. However, treatment for distant metastasis is warranted in cases of pancreatic adenosquamous carcinoma.

Characteristics of light chains of Chara myosin revealed by immunological investigation.

Chara myosin is plant myosin responsible for cytoplasmic streaming and moves actin filaments at 60 µm/s, which is the fastest of all myosins examined. The neck of the myosin molecule has usually mechanical and regulatory roles. The neck of Chara myosin is supposed to bind six light chains, but, at present, we have no knowledge about them. We found Ca⁺⁺-calmodulin activated Chara myosin motility and its actin-activated ATPase, and actually bound with the Chara myosin heavy chain, indicating calmodulin might be one of candidates for Chara myosin light chains. Antibody against essential light chain from Physarum myosin, and antibodies against Chara calmodulin and chicken myosin light chain from lens membranes reacted with 20 kDa and 18 kDa polypeptides of Chara myosin preparation, respectively. Correspondingly, column purified Chara myosin had light chains of 20 kDa, and 18 kDa with the molar ratio of 0.7 and 2.5 to the heavy chain, respectively.

Insight into the mechanism of fast movement of myosin from Chara corallina.

Chara myosin, two-headed plant myosin belonging to class XI, slides F-actin at maximally 60 microm s(-1). To elucidate the mechanism of this fast sliding, we extensively investigated its mechanochemical properties. The maximum actin activated ATPase activity, Vmax, was 21.3 s(-1) head(-1) in a solution, but when myosin was immobilized on the surface, its activity was 57.6 s(-1) head(-1) at 2 mg ml(-1) of F-actin. The sliding velocity and the actin activated ATPase activity were greatly inhibited by ADP, suggesting that ADP dissociation was the rate limiting step. With the extensive assay of motility by varying the surface density, the duty ratio of Chara myosin was found to be 0.49-0.44 from velocity measurements and 0.34 from the landing rate analysis. At the surface density of 10 molecules microm(-2), Chara myosin exhibited pivot movement under physiological conditions. Based on the results obtained, we will discuss the sliding mechanism of Chara myosin according to the working stroke model in terms of its physiological aspects. aspects.