• Title/Summary/Keyword: immunohistochemical methods

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KLK6 Promotes Growth, Migration, and Invasion of Gastric Cancer Cells

  • Zhu, Shengxing;Shi, Jihua;Zhang, Shanfeng;Li, Zhen
    • Journal of Gastric Cancer
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    • v.18 no.4
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    • pp.356-367
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    • 2018
  • Purpose: Kallikrein (KLK) proteases are hormone-like signaling molecules with critical functions in different cancers. This study investigated the expression of KLK6 in gastric cancer and its potential role in the growth, migration, and invasion of gastric cancer cells. Materials and Methods: In this study, we compared protein levels of KLK6, vascular endothelial growth factor (VEGF), and matrix metallopeptidase (MMP) 9 in normal gastric epithelial and gastric cancer cell lines by western blot. Fluorescence-activated cell sorting was employed to sort 2 clones of SGC-7901 cells with distinct KLK6 expression, namely, KLK6-high ($KLK6^{high}$) and KLK6-low ($KLK6^{low}$), which were then expanded. Lastly, immunohistochemical analysis was performed to investigate KLK6 expression in gastric cancer patients. Results: The expression levels of KLK6, VEGF, and MMP 9, were significantly higher in the gastric cancer cell lines SGC-7901, BGC-823, MKN-28, and MGC-803 than in the normal gastric epithelial cell line GES-1. Compared to $KLK6^{low}$ cells, $KLK6^{high}$ cells showed enhanced viability, colony-forming ability, migration, and invasion potential in vitro. Importantly, immunohistochemical analysis of a human gastric cancer tissue cohort revealed that the staining for KLK6, VEGF, and MMP9 was markedly stronger in the cancerous tissues than in the adjacent normal tissues. KLK6 expression also correlated with that of VEGF and MMP9 expression, as well as several key clinicopathological parameters. Conclusions: Together, these results suggest an important role for KLK6 in human gastric cancer progression.

Preventive Effect of Lactobacillus Fermentation Extract on Inflammation and Cytokine Production in Lipopolysaccharide-Induced Cystitis in Mice

  • Yoon, Hyun Suk;Kim, Yong Tae;Shim, Bong Suk;Yoon, Hana
    • Urogenital Tract Infection
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    • v.13 no.3
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    • pp.51-57
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    • 2018
  • Purpose: The effects of Lactobacillus fermentation extract (LFE) on cystitis induced by Escherichia coli lipopolysaccharide (LPS) in the mouse bladder were investigated by pathological analyses and measurement of the levels of tumor necrosis factor-alpha ($TNF-{\alpha}$) and interleukin-18 (IL-18). Materials and Methods: LFE was administered orally ($5{\mu}g/L$) to mice for 10 days after which the study group (n=12) received transurethral injection of $5{\mu}g/L$ LPS. The bladder tissue was then harvested after 24 hours and subjected to hematoxylin and eosin staining. A semi-quantitative score was used to evaluate inflammation (bladder inflammation index, BII). $TNF-{\alpha}$ immunohistochemical staining and multiplex cytokine assays were also performed. $TNF-{\alpha}$ and IL-18 levels were determined. The results were compared with those of the control group (n=12). Results: The BII in the control and study groups was $2.7{\pm}0.5$ and $1.1{\pm}0.7$, respectively, with the control group scores differing significantly from the study group scores (p<0.001). $TNF-{\alpha}$ immunohistochemical staining results were similar. The $TNF-{\alpha}$ levels determined by the multiplex cytokine assay were $2.82{\pm}1.35pg/mg$ and $1.55{\pm}0.56pg/mg$ for the control and study groups, respectively, and the difference between these groups was statistically significant (p=0.007). Conclusions: Oral administration of LFE appears to have a preventive effect against the inflammatory responses and $TNF-{\alpha}$ expression induced by transurethral instillation of LPS in the mouse bladder. Further studies are required to determine the clinical application of this finding.

Application of 630-nm and 850-nm Light-emitting Diodes and Microcurrent to Accelerate Collagen and Elastin Deposition in Porcine Skin

  • Kwon, Tae-Rin;Moon, Dong Wook;Kim, Jungwook;Kim, Hyoung Jun;Lee, Seong Jae;Han, Yunhee;Dan, Hee Won;Chi, Sang Hoon;Seong, Hwan Mo;Kim, Hee Jung;Lim, Guei-Sam;Lee, Jungkwan
    • Medical Lasers
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    • v.10 no.2
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    • pp.96-105
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    • 2021
  • Background and Objectives Skin aging is reportedly associated with regulation in collagen and elastin synthesis. This study investigated the potential of combining light-emitting diode (LED) treatments using a 630-nm and 850-nm LED with simultaneous microcurrent application. Materials and Methods The dorsal skin of female pigs was treated with a home-use device. We examined the treatment effects using photography, thermocamera, microscopic pathology, and histological examination to determine the mechanism of action, efficacy, and safety of the procedure. A histological observation was performed using hematoxylin and eosin, Masson's trichrome, Victoria blue, and immunohistochemical staining. We also used the Sircol soluble collagen and elastin assay kit to measure the amounts of collagen and elastin in the porcine back skin tissue after 2 and 6 weeks. Results Evaluation by visual inspection and devices showed no skin damage or heat-induced injury at the treatment site. Histological staining revealed that accurate treatment of the targeted dermis layer effectively enhanced collagen and elastin deposition. Collagen type I, a protein defined by immunohistochemical staining, was overexpressed in the early stages of weeks 2 and 6. Combined therapy findings showed the superior capability of the 630-nm and 850-nm LED procedures to induce collagen; in contrast, elastin induction was more pronounced after microcurrent treatments. Conclusion The home-use LED device, comprising a combination of 630-nm and 850-nm LEDs and microcurrent, is safe and can be used as an adjunctive treatment for self-administered facial rejuvenation.

Neurotropin protects rotator cuff tendon cells from lidocaine-induced cell death

  • Abe, Ryunosuke;Ohzono, Hiroki;Gotoh, Masafumi;Nakamura, Yosuke;Honda, Hirokazu;Nakamura, Hidehiro;Kume, Shinichiro;Okawa, Takahiro;Shiba, Naoto
    • Clinics in Shoulder and Elbow
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    • v.24 no.4
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    • pp.224-230
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    • 2021
  • Background: Local anesthetics often are used in rotator cuff tears as therapeutic tools, although some cases have reported that they have detrimental effects. Neurotropin (NTP) is used widely in Japan as a treatment for various chronic pain conditions and is shown to have protective effects on cartilage and nerve cells. In this study, we investigated the protective effect of NTP against lidocaine-induced cytotoxicity. Methods: Tenocytes from rotator cuff tendons were incubated with lidocaine, NTP, lidocaine with NTP, and a control medium. Cell viability was evaluated using the WST-8 assay. Cell apoptosis was detected via annexin V staining using flow cytometry. The expression of BCL-2 and cytochrome c, which are involved in the intrinsic mitochondrial pathway of apoptosis, was evaluated via Western blotting and immunohistochemical staining. Results: In the cell viability assay, lidocaine decreased cell viability in a dose-dependent manner, and NTP did not affect cell viability. Moreover, NTP significantly inhibited the cytotoxic effect of lidocaine. The flow cytometry analysis showed that lidocaine significantly induced apoptosis in tenocytes, and NTP considerably inhibited this lidocaine-induced apoptosis. Western blotting experiments showed that lidocaine decreased the protein expression of BCL-2, and that NTP conserved the expression of BCL-2, even when used with lidocaine. Immunohistochemical staining for cytochrome c showed that 0.1% lidocaine increased cytochrome c-positive cells, and NTP suppressed lidocaine-induced cytochrome c expression. Conclusions: NTP suppresses lidocaine-induced apoptosis of tenocytes by inhibiting the mitochondrial apoptotic pathway. Intra-articular/bursal injection of NTP with lidocaine could protect tenocytes in rotator cuff tendons against lidocaine-induced apoptosis.

Expression of Dopamine D2 Receptor in Response to Apomorphine Treatment in the Striatum of the Rat with Experimentally Induced Parkinsonism (파킨슨병 모형 흰쥐의 줄무늬체에서 Apomorphine 투여 방법에 따른 도파민 D2 수용체의 발현)

  • Choi, Seung Jin;Sung, Jae Hoon;Son, Byung Chul;Park, Choon Keun;Kwon, Sung Oh;Kim, Moon Chan;Lee, Sang Won
    • Journal of Korean Neurosurgical Society
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    • v.29 no.7
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    • pp.868-876
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    • 2000
  • Objective : Parkinsonian rat models have generally been characterized by unilateral destruction of both the nigrostriatal pathway and the mesolimbic pathway using the neurotoxin 6-hydroxydopamine. The induction of contraversive turning by apomorphine in these models is thought to reflect the stimulation of supersensitive dopamine D2 receptor or receptor-mediated mechanisms in denervated neostriatum. The present study was undertaken to investigate the expression of dopamine D2 receptor in denervated striatum according to modalities of apomorphine(dopamine agonist) treatment after creating a hemiparkinsonian rat model in which there is 6-hydroxydopamine induced destruction of the unilateral dopaminergic nigrostriatal pathway. Methods : After making complete lesion in left side substantia nigra pars compacta(SNpc) by stereotactic injection of 6-hydroxydopamine into medial and lateral areas of SNpc, and confirming successful animal model by apomorphine induced contraversive turning behavior without recovery and complete destruction of ipsilateral SNpc with tyrosine hydroxylase immunostaining in 7th day after operation, 15 rats of parkinsonian model were studied with or without administration of apomorphine at varying doses and durations. According to the modalities of apomorphine treatment for 4 days, these rats were divided into 3 groups, as not-treated group, intermittently treated group and constantly treated group. For investigating the extent of the expression of dopamine D2 receptor in denervated striatum, immunohistochemical staining by dopamine D2 receptor antibody and Western blot were performed. Results : In the D2 receptor antibody immunohistochemical staining, the mean number of positive stained neurons was highest in not-treated group($20.5{\pm}1.14$) of 3 groups. In constantly treated group, the mean number of positive stained neurons was less($3.9{\pm}1.79$) than intermittently treated group(p<0.05). The Western blotting with the D2 receptor antibody revealed that expression of receptors was also highest in not-treated group and less in constantiy treated group than intermittently treated group. Conclusion : Dopamine D2 receptors in denervated striatum of parkinsonian rat models, which were not treated with apomorphine, revealed to be most highly expressed. And, according to doses and durations of apomorphine administration, desensitization of the receptor was more apt to develop with constant treatment than intermittent treatment. In clinical setting, the authors believe that, in long-term treated parkinsonian patients, desensitization of dopamine receptors due to chronic dopaminergic stimulation seems to be partially related to mechanisms of drug tolerance.

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Evaluation of bone substitutes for treatment of peri-implant bone defects: biomechanical, histological, and immunohistochemical analyses in the rabbit tibia

  • dos Santos, Pamela Leticia;de Molon, Rafael Scaf;Queiroz, Thallita Pereira;Okamoto, Roberta;de Souza Faloni, Ana Paula;Gulinelli, Jessica Lemos;Luvizuto, Eloa Rodrigues;Garcia, Idelmo Rangel Junior
    • Journal of Periodontal and Implant Science
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    • v.46 no.3
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    • pp.176-196
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    • 2016
  • Purpose: We sought to evaluate the effectiveness of bone substitutes in circumferential periimplant defects created in the rabbit tibia. Methods: Thirty rabbits received 45 implants in their left and right tibia. A circumferential bone defect (6.1 mm in diameter/4 mm depth) was created in each rabbit tibia using a trephine bur. A dental implant ($4.1mm{\times}8.5mm$) was installed after the creation of the defect, providing a 2-mm gap. The bone defect gaps between the implant and the bone were randomly filled according to the following groups: blood clot (CO), particulate Bio-Oss$^{(R)}$ (BI), and Bio-Oss$^{(R)}$ Collagen (BC). Ten animals were euthanized after periods of 15, 30, and 60 days. Biomechanical analysis by means of the removal torque of the implants, as well as histologic and immunohistochemical analyses for protein expression of osteocalcin (OC), Runx2, OPG, RANKL, and TRAP were evaluated. Results: For biomechanics, BC showed a better biological response ($61.00{\pm}15.28Ncm$) than CO ($31.60{\pm}14.38Ncm$) at 30 days. Immunohistochemical analysis showed significantly different OC expression in CO and BC at 15 days, and also between the CO and BI groups, and between the CO and BC groups at 60 days. After 15 days, Runx2 expression was significantly different in the BI group compared to the CO and BC groups. RANKL expression was significantly different in the BI and CO groups and between the BI and BC groups at 15 days, and also between the BI and CO groups at 60 days. OPG expression was significantly higher at 60 days postoperatively in the BI group than the CO group. Conclusions: Collectively, our data indicate that, compared to CO and BI, BC offered better bone healing, which was characterized by greater RUNX2, OC, and OPG immunolabeling, and required greater reversal torque for implant removal. Indeed, along with BI, BC presents promising biomechanical and biological properties supporting its possible use in osteoconductive grafts for filling peri-implant gaps.

Correlation of Clinical Factors with HMGI(Y), p53 and Ki-67 Expression in Squamous Cell Carcinomas of the Head and Neck (두경부 편평세포암에서 HMGI(Y), p53, Ki-67의 발현과 임상인자와의 상관 관계)

  • Rho Young-Soo;Park Jun-Young;Park Il-Seok;Lim Young-Chang;Moon Sung-Ho;Kim Sung-Dong;Hwang Joon-Sik;Kim Duk-Hwan
    • Korean Journal of Head & Neck Oncology
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    • v.18 no.1
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    • pp.11-17
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    • 2002
  • Objectives: Expression of HMGI(Y), a nucleoprotein that binds to A/T rich sequences in the minor groove of the DNA helix, is observed in neoplastically transformed cells but not in normal cells. We have analyzed HMGI(Y), p53 expression and Ki-67 labelling index in squamous cell carcinomas of the head and neck, and evaluated its clinicopathologic significance. Materials and Methods: 40 cases of squamous cell carcinoma of the head and neck were entered on the study of immunohistochemical stains for HMGI(Y), p53 and Ki-67. We analyzed the relationship between HMGI(Y), p53, Ki-67 expression and age, sex, primary tumor site, stage, survival rate, recurrence. Results: HMGI(Y) expression evidenced by immunohistochemical staining was observed in 35 of 40 (87.5%) squamous cell carcinoma of the head and neck. But no significant correlation was observed between HMGI(Y) expression and other clinical factors such as primary site, tumor stage, differenciation, cervical lymph node, metastasis, recurrence and immunohistochemical status of p53. The Ki-67 labelling index was significantly correlated with recurrence and HMGI(Y) expression (p<0.05). Conclusion: This results suggest the Ki-67 is a good prognostic factor and the HMGI(Y) expression plays some roles in carcinogenesis and cellular proliferation of squamous cell carcinoma of the head and neck. HMGI(Y) gene can be used as a cancer marker, the correlation between the gene expression and the prognosis of the cancer patient should be proved in the future studies.

Correlations of Oral Tongue Cancer Invasion with Matrix Metalloproteinases(MMPs) and Vascular Endothelial Growth Factor(VEGF) Expression (Matrix Metalloproteinases(MMPs) 및 Vascular Endothelial Growth Factor(VEGF)의 발현을 통한 구강 설암의 침윤 기전 연구)

  • Kim Se-Heon;Cho Nam-Hoon;Lim Jae-Yul;Kim Ji-Hoon;Kim Jeong-Hong;Chang Jung-Hyun;Choi Eun-Chang
    • Korean Journal of Head & Neck Oncology
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    • v.21 no.1
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    • pp.3-9
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    • 2005
  • Purpose: In oral tongue cancer, the degree of tumor invasion has a significant effect on the prognosis. We hypothesized that the destruction of extracelluar matrix and neovascularization are related to tumor infiltration mechanism. By studying the the tissues of early stage oral tongue cancer patients, we are intend to clarify the invasion related factors in oral tongue cancer. Material and Methods: To demonstrate the invasion process in early T-stage oral tongue cancer, the expressions of extracellular matrix destruction related molecules(MMP2, MMP9) and neovascularization related molecule(VEGF) were observed by immunohistochemical study. Also, immunohistochemical staining of CD31 was done for quantification of neovascularization. With the experiment showed above, we analyzed relationship between expression of each substances and tumor invasion depth, tumor free survival rates and cervical lymph node metastasis rate in early T-stage oral tongue cancer. Results: The expression rates of MMP2, MMP9, VEGF in 38 early oral cancer patients were 52.6%, 78.9% 52.6%, respectively. Significant correlation was found between the VEGF expression and microvessel density showed by CD31 immunohistochemical staining(p<0.001). VEGF expressions were significantly related with tumor invasion depth(p=0.002). The tumor free survival rate of those patients with VEGF-positive tumors was significantly poorer than in those with VEGF-negative tumors(p=0.019). Conclusion: This results indicate that VEGF is a useful marker for predicting the tumor invasion in patients with early tongue cancer and could be used as a beneficial factors in defining operative field and prognosis.

Immunohistochemistry of Glucagon- immunoreactive Cells in the Developing Pancreas of the Korean Native Goat (Capra hircus)

  • Sae-Kwang Ku;Hyeung-Sik Lee;Jae-Hyun Lee
    • Animal cells and systems
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    • v.3 no.2
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    • pp.187-191
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    • 1999
  • The distribution of glucagon-immunoreactive cells in the pancreas during various developmental stages (fetus, neonate, 1-month-old, 6-month-old and adult) of the Korean native goat was investigated by immunohistochemical methods. The varying distribution and frequency of glucagon-immunoreactive cells in the pancreas of the Korean native goat were observed. The glucagon-immunoreactive cells were detected in both exocrine and endocrine portions (pancreatic islets) at all developmental stages and also in ducts of the 6-month-old and adult. The relative frequencies of glucagon-immunoreactive cells increased in the pancreatic islets and ducts with age, but decreased in the exocrine portions. Generally, they were distributed in the interacinar spaces or marginal zone of the pancreatic islets during all stages of development. However, the cell distributions of the pancreatic islets in the neonate divided into two types: 1) ones which were distributed in the inner zone, and 2) others in the peripheral zone.

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Congenital Dermatofibrosarcoma Protuberans: A Case Report (선천성 융기성 피부섬유육종: 증례보고)

  • Yun, Byung-Min
    • Archives of Plastic Surgery
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    • v.37 no.3
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    • pp.285-288
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    • 2010
  • Purpose: Dermatofibrosarcoma protuberans is a rare tumor, accounting for 0.1% of all malignant tumors. Although metastasis is very uncommon, local recurrence occur frequently. Dermatofibrosarcoma protuberans occurring in children is even more rare; this is the first case report of congenital dermatofibrosarcoma protuberans in Korea. Methods: The patient is a 14-month-old male infant with a lesion that was first thought to be a birthmark. The lesion grew larger, and a punch biopsy revealed dermatofibrosarcoma protuberans. A wide local excision was performed with a 2 cm peripheral resection margin beyond the gross tumor lesion. Deep fascia and a portion of muscle underneath the central part of the lesion were also taken. The surgical defect was covered by a split-thickness skin graft. Results: There has been no clinical sign of recurrence over one year after the surgery. Conclusion: A patient with congenital dermatofibrosarcoma protuberans detected at an early stage underwent a wide local excision of the tumor after accurate diagnosis was carried out by biopsy and immunohistochemical studies. There was no clinical evidence of tumor recurrence during over a 1-year follow-up.