Dense distribution of macrophages in flexor aspects of the hand and foot of mid-term human fetuses / 대한해부학회지

In the developing human musculoskeletal system, cell death with macrophage accumulation occurs in the thigh muscle and interdigital area. To comprehensively clarify the distribution of macrophages, we immunohistochemically examined 16 pairs of upper and lower extremities without the hip joint (left and right sides) obtained from 8 human fetuses at approximately 10-15 weeks of gestation. Rather than in muscles, CD68-positive macrophages were densely distributed in loose connective tissues of the flexor aspects of the extremities, especially in the wrist, hand and foot. In contrast, no or fewer macrophages were evident in the shoulder and the extensor aspects of the extremities. The macrophages were not concentrated at the enthesis of the tendon and ligament, but tended to be arranged along other connective tissue fibers. Deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling revealed apoptosis in the hand lumbricalis muscles, but not in the area of macrophage accumulation. Likewise, podoplanin-positive lymphatic vessels were not localized to areas of macrophage accumulation. Re-organization of the connective tissue along and around the flexor tendons of the hand and foot, such as development of the bursa or tendon sheath at 10-15 weeks, might require the phagocytotic function of macrophages, although details of the mechanism remain unknown.

Neuroprotective effects of L-carnitine against oxygen-glucose deprivation in rat primary cortical neurons / 소아과

PURPOSE: Hypoxic-ischemic encephalopathy is an important cause of neonatal mortality, as this brain injury disrupts normal mitochondrial respiratory activity. Carnitine plays an essential role in mitochondrial fatty acid transport and modulates excess acyl coenzyme A levels. In this study, we investigated whether treatment of primary cultures of rat cortical neurons with L-carnitine was able to prevent neurotoxicity resulting from oxygen-glucose deprivation (OGD). METHODS: Cortical neurons were prepared from Sprague-Dawley rat embryos. L-Carnitine was applied to cultures just prior to OGD and subsequent reoxygenation. The numbers of cells that stained with acridine orange (AO) and propidium iodide (PI) were counted, and lactate dehydrogenase (LDH) activity and reactive oxygen species (ROS) levels were measured. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and the terminal uridine deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling assay were performed to evaluate the effect of L-carnitine (1 microM, 10 microM, and 100 microM) on OGD-induced neurotoxicity. RESULTS: Treatment of primary cultures of rat cortical neurons with L-carnitine significantly reduced cell necrosis and prevented apoptosis after OGD. L-Carnitine application significantly reduced the number of cells that died, as assessed by the PI/AO ratio, and also reduced ROS release in the OGD groups treated with 10 microM and 100 microM of L-carnitine compared with the untreated OGD group (P<0.05). The application of L-carnitine at 100 microM significantly decreased cytotoxicity, LDH release, and inhibited apoptosis compared to the untreated OGD group (P<0.05). CONCLUSION: L-Carnitine has neuroprotective benefits against OGD in rat primary cortical neurons in vitro.

Establishment of a hydrogel chip for high-throughput detection of Y chromosome microdeletions / 中华男科学杂志

<p><b>OBJECTIVE</b>To establish a high-sensitivity, high-specificity and low-cost hydrogel chip platform for the clinical screening of Y chromosome microdeletions.</p><p><b>METHODS</b>Site-specific extended primers with a common sequence at the 5' end were used for hybridizing with the target. The Cy5-dUTP was incorporated into the products by primer extension, and the products were labeled with fluorescence. Then the extended products were added to the chip for hybridizing with acrylamide-modified common probes immobilized on the chip. After removal of the free Cy5-dUTP by electrophoresis, the signals were obtained by fluorescence scanning. And the detecting conditions of this method were optimized.</p><p><b>RESULTS</b>SY254 of 9 samples was successfully detected with the hydrogel chip. The results showed that 3 were normal and the other 6 with microdeletions (1 female sample as a negative control), which coincided with the results of conventional multiplex PCR-electrophoresis.</p><p><b>CONCLUSION</b>The hydrogel chip platform we established has provided a new technique for the detection of Y chromosome microdeletions, and is beneficial to the diagnosis and treatment of male infertility.</p>

Is It Possible to Recover Erectile Function Spontaneously after Cavernous Nerve Injury? Time-Dependent Structural and Functional Changes in Corpus Cavernosum Following Cavernous Nerve Injury in Rats / 대한남성과학회지

PURPOSE: There has been a scarcity of integrated, long-term (>4 week) studies on structural and functional alterations in the penis according to the period following cavernous nerve (CN) injury. The aim of this study was to investigate time-dependent structural and functional changes in the corpus cavernosum following CN injury in a rat model. MATERIALS AND METHODS: Ninety male Sprague-Dawley rats (10 weeks old) were divided into 4 groups: normal control (C), sham (S), bilateral CN resection (R), and bilateral CN crush injury (I) groups. At 1, 4, and 12 weeks after the procedure, erectile function was assessed by electrostimulation. The terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine 5'-triphosphate nick end labeling (TUNEL) assay was performed for detection of apoptosis. Masson's trichrome staining and immunohistochemistry were performed for detection of alpha smooth muscle actin (alpha-SMA). Western blot analysis was then performed. RESULTS: The R and I groups showed persistent impairment of erectile function at all three points in time. Apoptosis peaked at 1 week after resection or crush injury and then gradually subsided. The smooth muscle cell/collagen ratio and expression of alpha-SMA gradually decreased over time after CN resection or crush injury. Myosin phosphatase target subunit 1 phosphorylation progressively increased over time after CN resection or crush injury. On the other hand, expression of phospho-protein kinase B, phospho-endothelial nitric oxide synthase, and neuronal nitric oxide synthase transiently decreased at 1 week after resection or crush injury and then recovered to the control values. CONCLUSIONS: Our results suggest that persistent up-regulation of the RhoA/Rho-kinase pathway and structural change such as decreased smooth muscle cell and increased cavernosal fibrosis might play an important role in persistent erectile dysfunction following CN injury.

Localization of herpes simplex virus type 1 DNA in latently infected BALB/c mice neurons using in situ polymerase chain reaction

Herpes simplex virus type-1 [HSV-1] establishes a lifelong latent infection in neurons following primary infection. The existence of latent HSV-1 DNA in the trigeminal ganglia of infected BALB/c mice was examined using a direct in situ PCR technique, based on Digoxigenin-11-dUTP detection system with anti-digoxigenin-peroxidase and 3,3f-diaminobenzidine [DAB] substrate. Eight-week-old male BALB/c mice were inoculated via the eye by 10[4] plaque forming unit of wild type Iranian isolates of HSV-1. After establishment of latency, trigeminal ganglia were removed and examined using in situ PCR to detect HSV-1 genome. Finally, the results of in situ PCR were verified by a two-round PCR method, using amplification cocktail of in situ reaction, as a template for a conventional gel base PCR. The results suggest that a direct in situ PCR method using a peroxidase and DAB detection system is a useful means for detection of latent HSV-1 DNA in the latently infected ganglia

The Clinical Features and Pathophysiology of Acute Radiation Dermatitis in Patients Receiving Tomotherapy

BACKGROUND: Radiation therapy (RT) including tomotherapy has been widely used to treat primary tumors, as well as to alleviate the symptoms of metastatic cancers. OBJECTIVE: The primary purpose of this study was to examine the characteristics of the clinical features and pathophysiological mechanisms associated with acute radiation dermatitis in cancer patients that received tomotherapy, and compare the results to patients treated by conventional radiation therapy. METHODS: The study population consisted of 11 patients that were referred to the dermatology department because of radiation dermatitis after receiving tomotherapy; all patients were evaluated for clinical severity. The patients were assessed and identified using the National Cancer Institute Common Toxicity Criteria version (CTC) 3.0. We performed biopsies of the skin lesions that were examined for apoptosis using the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labelling (TUNEL) assay and stained immunohistochemically with monoclonal antibodies to CD8, CD4 and TGF-beta. As a positive control, patients with radiation dermatitis treated with conventional radiation therapy were also studied. RESULTS: The results of the clinical features of the skin of tomotherapy patients were the following: grade 1 (36%), grade 2 (55%) and other changes (9%). Among the population that had skin lesions due to acute radiation dermatitis, the mean number of positive cells per high power field (HPF) was the following: there were 30.50+/-.50 TUNEL- positive cells, 34.60+/-12.50 CD8+ T cells, 5.19+/-3.17 CD4+T cells and 9.95+/-1.33 TGF-beta positive cells measured per HPF. The mean number of positive cells per HPF for the patients that received conventional radiation therapy was: TUNLEL-positive cells in 7.5+/-1.64, CD8-, CD4- and TGF-beta-positive cells in 12.50+/-3.73, 3.16+/- 1.47, 6.50+/-1.97. CONCLUSION: We found that the number of TUNEL-positive cells and CD8+ T cells were higher in the lesions of patients receiving tomotherapy compared to the lesions of the patients receiving conventional radiation therapy. These findings suggest that tomotherapy without dose modification may cause significantly more severe forms of radiation dermatitis by apoptosis and cytotoxic immune responses than conventional radiation therapy.

Expression of Survivin in Gastric Carcinoma and its Relation to Tumor Cell Proliferation and Apoptosis

BACKGROUND: Survivin, a novel antiapoptotic gene has been linked with tumor development and progression in various human carcinomas including gastric carcinomas. The aim of this study was to evaluate the expression of survivin in gastric carcinoma and its correlation with tumor cell proliferation and apoptosis. METHODS: Expression of survivin was evaluated immunohistochemically in 84 surgically resected gastric carcinomas. Tumor cell apoptosis was evaluated with terminal deoxynucleotidyl transferase (TdT) mediated deoxyuridine triphosphate (dUTP) nick-end labeling (TUNEL), and Ki-67 immunostaining was used for evaluation of tumor cell proliferation. RESULTS: Expression of survivin was noted in 53.6% of the gastric carcinomas, and was significantly associated with depth of invasion, status of lymph node metastasis or tumor stage (p=0.022, 0.034, 0.040, respectively). There was an inverse correlation between survivin expression and apoptotic index (p=0.015). But there was no significant correlation between survivin expression and Ki-67 labeling index (p=0.430). CONCLUSIONS: These results suggest that survivin expression may contribute to tumor development and progression by inhibiting apoptosis in human gastric carcinoma.

Affection of metallothionein-3 to dUTPase's accommodating cellular toxicity of dUTP / 生物工程学报

Metallothionein-3 (MT-3), renamed as growth inhibitory factor (GIF), is a brain specific member of the metallothionein family. Human dUTPase is a recently found protein in brain that can interact with hMT-3. They have the growth inhibitory activity on neuron cell by interaction. To study the affection of hMT-3 to dUTPase's eliminating the cellular toxicity caused by dUTP, the pSVHA-dUTPase and pFLag-hMT-3 genes have been transfected into HEK293 cells. In addition, the dUTPase and hMT-3 proteins were expressed in BL21 to study the role of hMT-3 on the hydrolyzation of dUTP by dUTPase. The results demonstrate that the cells co-transfected with dUTPase and hMT-3 genes have more strong resistibility to dUTP than the cells transfected only with dUTPase gene. And that the hMT-3 protein can accelerate the hydrolyzation of dUTP by dUTPase. All these indicate that hMT-3 can cooperate with dUTPase to protect better the 293 cells from dUTP. This research offered the theoretic elements for the application of hMT-3 and dUTPase in chemic cure.