Matrigel‐Based Sprouting Endothelial Cell Culture System from Mouse Corpus Cavernosum Is Potentially Useful for the Study of Endothelial and Erectile Dysfunction Related to High‐Glucose Exposure

IntroductionA proper cavernous endothelial cell culture system would be advantageous for the study of the pathophysiologic mechanisms involved in endothelial dysfunction and erectile dysfunction (ED).AimTo establish a nonenzymatic technique, which we termed the “Matrigel‐based sprouting endothelial cell culture system,” for the isolation of mouse cavernous endothelial cells (MCECs) and an in vitro model that mimics in vivo situation for diabetes‐induced ED.MethodsFor primary MCEC culture, mouse cavernous tissue was implanted into Matrigel and sprouting cells from the tissue were subcultivated. To establish an in vitro model for diabetes‐induced ED, the primary cultured MCECs were exposed to a normal‐glucose (5 mmoL) or a high‐glucose (30 mmoL) condition for 48 hours.Main Outcome MeasuresThe purity of isolated cells was determined by fluorescence‐activated cell sorting analysis. MCECs incubated under the normal‐ or the high‐glucose condition were used for Western blot, cyclic guanosine monophosphate (cGMP) quantification, and in vitro angiogenesis assay.ResultsWe could consistently isolate high‐purity MCECs (about 97%) with the Matrigel‐based sprouting endothelial cell culture system. MCECs were subcultured up to the fifth passage and no significant changes were noted in endothelial cell morphology or purity. The phosphorylation of Akt and eNOS and the cGMP concentration were significantly lower in MCECs exposed to high glucose than in those exposed to normal glucose. MCECs exposed to the normal‐glucose condition formed well‐organized capillary‐like structures, whereas derangements in tube formation were noted in MCECs exposed to high glucose. The protein expression of transforming growth factor‐β1 (TGF‐β1) and phospho‐Smad2 was significantly increased by exposure to high glucose.ConclusionThe Matrigel‐based sprouting endothelial cell culture system is a simple, technically feasible, and reproducible technique for isolating pure cavernous endothelial cells in mice. An in vitro model for diabetic ED will be a valuable tool for evaluating the angiogenic potential of novel endogenous or synthetic modulators. Yin GN, Ryu J‐K, Kwon M‐H, Shin SH, Jin HR, Song K‐M, Choi MJ, Kang D‐Y, Kim WJ, and Suh J‐K. Matrigel‐based sprouting endothelial cell culture system from mouse corpus cavernosum is potentially useful for the study of endothelial and erectile dysfunction related to high‐glucose exposure. J Sex Med 2012;9:1777–1789.     

Heterogeneity of cancer‐associated fibroblasts: Opportunities for precision medicine

Despite marked development in cancer therapies during recent decades, the prognosis for advanced cancer remains poor. The conventional tumor–cell‐centric view of cancer can only explain part of cancer progression, and thus a thorough understanding of the tumor microenvironment (TME) is crucial. Among cells within the TME, cancer‐associated fibroblasts (CAFs) are attracting attention as a target for cancer therapy. However, CAFs present a heterogeneous population of cells and more detailed classification of CAFs and investigation of functions of each subset is needed to develop novel CAF‐targeted therapies. In this context, application of newly developed approaches to single‐cell analysis has already made an impact on our understanding of the heterogeneity of CAFs. Here, we review the recent literature on CAF heterogeneity and function, and discuss the possibility of novel therapies targeting CAF subsets.     

Dysgerminoma of the ovary with precocious puberty: A case report

Ovarian dysgerminoma is usually hormonally inert, but when it contains syncytiotrophoblastic giant cells or undergoes malignant transformation, the level of estradiol might be elevated. A 6-year-old girl contracted ovarian dysgerminoma with precocious puberty, and her levels of β-human chorionic gondaotropin, α-fetoprotein and estradiol were high. After resection of the tumor, levels of tumor markers became normal and precocious puberty disappeared. We report this unusual type of dysgerminoma with a brief review of the literature.     

Diagnostic yield of endoscopic ultrasonography-guided single-incision needle knife biopsy for gastric subepithelial tumors: comparison with resected specimens

Background

Several techniques are recommended for the histologic diagnosis of gastric subepithelial tumors (SETs). The purpose of our study was to evaluate the diagnostic yield and safety of endoscopic ultrasonography-guided single-incision needle knife (SINK) biopsy for the diagnosis of gastric SETs.

Methods

A retrospective review of patients who received biopsy for gastric SETs from August 2012 to May 2015 was conducted. Patients who received endoscopic ultrasonography and were found to have a SET originating from the muscularis propria of the stomach were included in the study. The aim of our study was to investigate the safety and diagnostic yield of SINK biopsy for gastric SETs.

Results

A total of 31 patients received SINK biopsy for SETs. The diagnostic yield of SINK biopsy was 87 % (95 % CI 75–100 %), and the diagnostic accuracy was 89 % (95 % CI 74–105 %). The sensitivity of SINK biopsy to identify gastrointestinal stromal tumors was 83 % (95 % CI 52–98 %); the specificity was 100 % (95 % CI 59–100 %); the positive predictive value was 100 % (95 % CI 69–100 %); and the negative predictive value was 78 % (95 % CI 40–97 %). There were no procedure-related adverse events during and after procedure.

Conclusion

The use of SINK biopsy technique in patients with SETs is a good diagnostic tool with high diagnostic yield and accuracy. The method is simple, safe, and associated with few complications.

     

Image quality assessment of ultra low-dose chest CT using sinogram-affirmed iterative reconstruction

Objectives

To assess the image quality of ultra-low-dose computed tomography (ULDCT) using sinogram-affirmed iterative reconstruction (SAFIRE) compared to reduced dose CT (RDCT).

Methods

Eighty-one consecutive patients underwent non-enhanced ULDCT using 80 kVp and 30 mAs and contrast-enhanced RDCT using automated tube potential selection and tube current modulation. CT images were reconstructed with SAFIRE. Image noise and subjective image quality of normal structures and various pulmonary lesions were assessed.

Results

The mean effective doses were 0.29?±?0.03 and 2.88?±?1.11 mSv for ULDCT and RDCT, respectively. ULDCT had significantly higher noise (p?<?0.001). Image quality of five normal structures was diagnostic in 91.1 % of ULDCT and 100 % of RDCT. With ULDCT, the frequencies of non-diagnostic image quality were 2.0 (1/50), 4.6 (13/280), 25.5 (14/55), and 40.0 (8/20)% for BMIs of?<?20, 20–25, 25–30, and >30. In the assessment of pulmonary lesions, non-diagnostic image quality was observed for 11.2 % of all lesions, 60.9 % of decreased attenuation (significantly more frequent for upper lung lesions), and 23.5 % of ground-glass nodules.

Conclusion

ULDCT generates diagnostic images in patients with a BMI ≤25, but is of limited use for lesions with decreased attenuation, ground-glass nodules, or those located in the upper lobe.

Key Points

? Iterative reconstruction enables ultra-low-dose CT (ULDCT) with very low radiation doses. ? Image quality of ULDCT depends on the patient body mass index (BMI). ? Selection of kVp and mAs depends on both BMI and lesion type. ? Diagnosis of pulmonary emphysema or ground-glass nodules requires higher radiation doses.     

Visceral obesity is a better predictor than generalized obesity for basal insulin requirement at the initiation of insulin therapy in patients with type 2 diabetes

Aims

Basal insulin requirement in patients with type 2 diabetes is difficult to determine because of individual variability in insulin sensitivity and secretion. We aimed to identify factors that influence basal insulin requirement in insulin-naïve patients with type 2 diabetes.

Methods

We studied 50 insulin-naïve patients with type 2 diabetes. Their basal insulin requirement was calculated by 8 h overnight intravenous insulin infusion. Patients underwent abdominal computed tomography; subcutaneous and visceral fat areas were measured.

Results

The basal insulin requirement was 31.3 ± 16.9 units/day, and it varied widely from 0.2 to 1.4 units/kg. It was positively correlated with visceral fat area (γ = 0.485, P < 0.001), body mass index (BMI, γ = 0.339, P = 0.008), glycated hemoglobin (HbA1C, γ = 0.327, P = 0.019), alanine aminotransferase (ALT, γ = 0.310, P = 0.027), and triglyceride (γ = 0.305, P = 0.032). However, body weight, waist circumference and total fat mass were not related to basal insulin requirement. Multiple linear regression analysis showed that visceral fat area, HbA1C, and ALT are independent predictors of basal insulin requirement.

Conclusions

Visceral obesity was a better predictor than generalized obesity for basal insulin requirement at the initiation of insulin therapy in patients with type 2 diabetes.     

Clinicopathologic Characteristics of Breast Cancer Stem Cells Identified on the Basis of Aldehyde Dehydrogenase 1 Expression

Purpose

Breast cancer displays varying molecular and clinical features. The ability to form breast tumors has been shown by several studies with aldehyde dehydrogenase 1 (ALDH1) positive cells. The aim of this study is to investigate the association between ALDH1 expression and clinicopathologic characteristics of invasive ductal carcinoma.

Methods

We investigated breast cancer tissues for the prevalence of ALDH1⁺ tumor cells and their prognostic value. The present study included paraffin-embedded tissues of 70 patients with or without recurrences. We applied immunohistochemical staining for the detection of ALDH1⁺ cells. Analysis of the association of clinical outcomes and molecular subtype with marker status was conducted.

Results

ALDH1⁺ and ALDH1^- tumors were more frequent in triple-negative breast cancers and in luminal A breast cancers, respectively (p<0.01). ALDH1 expression was found to exert significant impact on disease free survival (DFS) (ALDH1⁺ vs. ALDH1^-, 53.1±6.7 months vs. 79.2±4.7 months; p=0.03) and overall survival (OS) (ALDH1⁺ vs. ALDH1^-, 68.5±4.7 months vs. 95.3±1.1 months; p<0.01). In triple-negative breast cancer (TNBC) patients, DFS and OS showed no statistical differences according to ALDH1 expression (ALDH1⁺ vs. ALDH1^-, 45.3±9.4 months vs. 81.3±7.4 months, p=0.52; 69.0±7.5 months vs. 91.3±6.3 months, p=0.67). However, non-TNBC patients showed significant OS difference between ALDH1⁺ and ALDH1^- tumors (ALDH1⁺ vs. ALDH1^-, 77.6±3.6 months vs. 98.0±1.0 months; p=0.04) with no statistical difference of DFS (ALDH1⁺ vs. ALDH1^-, 60.5±8.0 months vs. 81.8±4.6 months; p=0.27).

Conclusion

Our findings suggest that the expression of ALDH1 in breast cancer may be associated with TNBC and poor clinical outcomes. On the basis of our findings, we propose that ALDH1 expression in breast cancer could be correlated with poor prognosis, and may contribute to a more aggressive cancer phenotype.     

Substance P augments a persistent slow inward calcium-sensitive current in voltage-clamped spinal dorsal horn neurons of the rat

Rat spinal dorsal horn neurons in slice preparations perfused with Ringer solution containing 0.5-1 microM TTX and/or 10-20 mM tetraethylammonium at 29 degrees C, were studied by using a single microelectrode voltage-clamp technique. Slow persistent inward currents were recorded during depolarizing voltage commands to membrane potentials positive to about -40 mV. The inward current was depressed by removing external Ca, or by adding 0.1-0.2 mM Cd, 5 mM Co or 0.1 mM verapamil, and was increased by adding Ba or Bay-K 8644. Substance P (SP) augmented a persistent slow inward Ca-sensitive current in a dose-dependent manner. It is suggested that this effect may be instrumental in generating the SP-evoked slow depolarization, increase in membrane excitability, and the 'bursting' behavior in the immature rat dorsal horn neurons. In addition, in some neurons SP reduced the M-like current, which effect may contribute to, but not explain, generation of the SP-induced slow depolarization.     

Hepatic resection using a liver-hanging maneuver and Glissonean pedicle transection for hepatocellular carcinoma in a patient with situs inversus totalis: report of a case

Situs inversus totalis is a rare congenital anomaly in which the major abdominal organs are located as a mirror image of their normal positions. This poses much difficulty for surgeons. We describe how we performed the liver-hanging maneuver (LHM) for hepatocellular carcinoma (HCC) in a 59-year-old man with situs inversus totalis, to resolve the difficulty of the mirror-image location of his liver. The HCC was located in the right lateral sector. Although segmentectomy of segment 7 would normally be considered minimal for a curative treatment of HCC, this was relatively complicated in this patient. Thus, we performed an extended right lateral sectionectomy using the LHM to achieve a simple transection. The hepatic hilum was dissected using the Glissonean pedicle transection method. The operation time and intraoperative blood loss were 6 h 45 min and 471 ml, respectively. No blood product transfusion was required. The LHM and the hilar Glissonean pedicle approach proved effective for resolving the difficulties of performing surgery in a mirror image for HCC in a patient with situs inversus totalis.