immunohistochemistry staining
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2022 ◽  
Author(s):  
Shimao Song ◽  
Xinzhou Deng ◽  
Jincheng Wang ◽  
Jiahui Han ◽  
Zhen Peng ◽  
...  

Abstract Background: GRAP2 is an adaptor protein involved in leukocyte-specific protein-tyrosine kinase signaling; however, the prognostic value of GRAP2 and its correlation with immune cell infiltration in lung adenocarcinoma (LUAD) remain unclear.Methods: All original data were downloaded from the TCGA database and integrated via R 3.2.2. GRAP2 expression was explored with the TCGA and TIMER databases. We evaluated the influence of GRAP2 on clinical prognosis using the Kaplan-Meier plotter, Gene Expression Omnibus (GEO) database and GEPIA database. Correlations between GRAP2 and cancer immune characteristics were analyzed via TIMER and TISIDB databases. Finally, we confirmed the expression of GRAP2 in LUAD by immunohistochemistry staining.Results: Transcription levels of GRAP2 were significantly lower in several human cancers, including LUAD, than in adjacent normal tissues. We also found that tumor tissues have lower protein expression levels of GRAP2 compared with adjacent normal tissues in LUAD by immunohistochemistry staining. The down-regulated GRAP2 was associated with poorer overall survival, pathologic stage, T stage, N stage and primary therapy outcome in LUAD. Mechanically, we identified a hub gene that included a total of 91 GRAP2 co-expressed genes, which were tightly associated with immune response in LUAD. GRAP2 expression was positively correlated with infiltrating levels of B cells, CD8+ T cells, dendritic cells, eosinophils, macrophages, mast cells, Th2 cells, Th1 cells, Th17 cells, NK cells and neutrophils. GRAP2 expression level also affected the cumulative survival time of B cells and dendritic cells. GRAP2 expression is positively correlated with multiple immune markers, chemokines, chemokine receptors and MHC molecules of LUAD.Conclusions: These findings suggest that GRAP2 is a tumor suppressor gene and can be used as a prognostic biomarker for determining prognosis and immune infiltration in LUAD.


2021 ◽  
Author(s):  
Nanni Hua ◽  
Anxian Chen ◽  
Chen Yang ◽  
Hui Dong ◽  
Xianglei He ◽  
...  

Abstract Fibrinogen-like protein-1 (FGL1), as the member of FREP superfamily, had been widely concerned as a major immune inhibitory ligand of LAG-3. Although FGL1 expression levels didn’t have significant difference in most of tumors via online data analysis, we found that it was down-regulated in liver cancer. Moreover, the correlation between FGL1 expression and the progression and prognosis of hepatocellular carcinoma (HCC) was still disputed. In our study, firstly, we used bioinformatics analysis to define the expression profile and clinical significance of FGL1 in HCC. Then, we determined the FGL1 level in 9 human HCC cell lines and 11 pairs of tumor and normal liver tissues from HCC patients by western blot. Furthermore, tissue microassays were used to detect the expression of FGL1 through immunohistochemistry staining and verify whether FGL1 expression levels were associated with clinicopathological features of HCC patients or not. The results of the experiment proved that FGL1 was down-regulated significantly in HCC cell lines and HCC tissues, corresponding with the results of our bioinformatics analysis. FGL1 expression levels in HCC were related to Edmondson grade and metastasis. Additionally, high FGL1 expression was related to better overall survival in HCC patients, indicating that the down-regulated FGL1 was correlated with poor prognosis and FGL1 might function as a tumor suppressor. Taken together, expression levels of FGL1 may correlate with the progression and prognosis of HCC, and FGL1 could be a potential prognostic biomarker.


2021 ◽  
Vol 10 (24) ◽  
pp. 5896
Author(s):  
Simona R. Gheorghe ◽  
Cees Vermeer ◽  
Gabriel Olteanu ◽  
Ciprian N. Silaghi ◽  
Alexandra M. Crăciun

Matrix Gla protein (MGP), a local inhibitor of tissue mineralization, is associated with vascular calcification. Depending on the carboxylation and phosphorylation status, MGP has active conformations, e.g., carboxylated MGP (cMGP) and phosphorylated MGP (pMGP), but also inactive conformations, e.g., uncarboxylated MGP (ucMGP) and dephosphorylated MGP (dpMGP). Our purpose was to assess the presence of all MGP conformations in healthy veins (HV) and varicose veins (VV), concurrently with the analysis of circulating total MGP (tMGP) before and after the surgical stripping of VV. We collected samples from the great saphenous vein, considered as control group, and tissue from VV, designated as VV group. Plasma levels of tMGP were significantly decreased after the surgical removal of the VV (before 59.5 ± 17.2 vs. after 38.1 ± 11.3, p < 0.001). By using immunohistochemistry staining, we identified local cMGP and pMGP in the control and VV groups, both without calcification, while ucMGP and dpMGP were absent. cMGP was observed in the nucleus and cytoplasm and pMGP in the nucleus of cells belonging to the tunica media, tunica intima and vasa vasorum. Therefore, the active conformations of MGP (cMGP and pMGP) are prevalent in HV and VV without calcification, affirming their anti-calcifying role in veins.


2021 ◽  
Vol 2021 ◽  
pp. 1-10
Author(s):  
Peng Liu ◽  
Chen Wang ◽  
Yun Wang ◽  
Honghong Zhang ◽  
Baoli Liu ◽  
...  

Hyperuricemia, an independent risk factor for ensuing chronic kidney disease (CKD), contributed to tubulointerstitial fibrosis and insufficiency of renal fatty acid oxidation. Many studies have shown that renal fatty acid oxidation dysfunction is related to the TGF-β1/Smad3 signaling pathway. We experimented the effects of Zishen Qingre Tongluo Formula (ZQTF) on the adenine/yeast-induced HN rats and uric acid-induced renal mouse tubular epithelial cells (mTECs), determined whether this effect was related to the TGF-β1/Smad3 signaling pathway, and further investigated the relationship between this effect and renal fatty acid oxidation. Rats were given intraperitoneally with adenine (100 mg/kg) and feed chow with 10% yeast for 18 days and then received ZQTF (12.04 g/kg/day) via intragastric gavage for eight weeks. The TGF-β1/Smad3 signaling pathway and renal fatty acid oxidation protein were detected by using western blotting, real-time PCR, and immunohistochemistry staining. mTECs induced by UA were used to investigate the relationship between the TGF-β1/Smad3 signaling pathway and renal fatty acid oxidation. After treatment with ZQTF, levels of UA, 24 h UTP, BUN, and Scr were significantly decreased and histologic injuries were visibly ameliorated in HN rats. Western blotting, real-time PCR, and immunohistochemistry staining revealed that PGC-1α, PPARγ, and PPARα significantly increased, and fibronectin, collagen 1, and P-Smad3 significantly decreased in HN rats and UA-induced mTECs after ZQTF treatment. SIS3 (a specific inhibitor of Smad3) treatment significantly increased the expressions of PGC-1α, PPARγ, and PPARα and decreased the expressions of fibronectin, collagen 1, and P-Smad3 in UA-induced mTECs. Our study demonstrated that ZQTF exhibited renoprotective effects by promoting renal fatty acid oxidation via the regulation of the TGF-β1/Smad3 signaling pathway. Thus, the present results indicated that ZQTF was a novel antifibrotic strategy for hyperuricemic nephropathy.


2021 ◽  
Vol 2021 ◽  
pp. 1-9
Author(s):  
Chia-Li Chung ◽  
Hung-Pei Tsai ◽  
Yu-Hua Huang ◽  
Shu-Chuan Wu ◽  
Chee-Yin Chai ◽  
...  

Background. Subarachnoid hemorrhage (SAH) is an important subcategory of stroke due to its high mortality rate as well as severe complications such as neurological deficit. It has been suggested that cerebral inflammation is a major factor in advanced brain injury after SAH. Microglia and astrocytes are known supporting cells in the development and maintenance of inflammation in central nervous system. However, the role of microglia and astrocytes in the development of inflammation and neuronal cell apoptosis during the early phase after SAH has not been thoroughly investigated. Materials and Methods. Sprague-Dawley rats were divided into 4 groups ( n = 6 /group): sham group, animals subjected to SAH without treatment, SAH animals pretreated with the microglia inhibitor minocycline (50 mg/kg, ip), and SAH animals pretreated with the astrocyte inhibitor fluorocitrate (50 mg/kg, ip). SAH was induced by injecting autologous blood (1 ml/kg) into the cistern magna on day 0. Pretreatment with minocycline or fluorocitrate was given three days prior to the induction of SAH. Rats were sacrificed 6 hr after SAH, and their cerebral spinal fluids were used to measure protein levels of neuroinflammatory cytokines IL-1β, IL-6, and TNF-α by ELISA. In addition, the cerebral cortex was utilized to determine the levels of caspase-3 by western blot and to evaluate neuronal cell apoptosis by immunohistochemistry staining and detect microglia and astrocyte by immunofluorescence staining for Iba-1 and GFAP. In this study, all SAH animals were given an injection of autologous blood and SAH rats treated with minocycline or fluorocitrate received ip injections on day 1, 2, and 3 before inducing SAH. Neurological outcome was assessed by ambulation and placing/stepping reflex responses on day 7. Results. Immunofluorescence staining showed that SAH induced proliferation of microglia and astrocyte and minocycline inhibited the proliferation of both microglia and astrocyte. However, fluorocitrate inhibited only the proliferation of astrocyte. ELISA analysis showed that SAH upregulated TNF-α and IL-1β, but not IL-6 at 6 hr after SAH. Minocycline, but not fluorocitrate, attenuated the upregulation of TNF-α and IL-1β. Western blot analysis and immunohistochemistry staining showed that SAH induced neuronal cell apoptosis. Pretreatment with minocycline, but not fluorocitrate, decreased SAH-induced neuronal death and cerebral vasospasm. Furthermore, significant improvements in neurobehavioral outcome were seen in the minocycline treatment group, but not in animals treated with fluorocitrate. Conclusions. Microglia may play an important role to regulate neuronal cell apoptosis and cerebral vasospasm through inhibiting inflammation at an early phase after SAH in the rat.


Author(s):  
Nina Struve ◽  
Konstantin Hoffer ◽  
Anna-Sophie Weik ◽  
Britta Riepen ◽  
Leonie Krug ◽  
...  

Abstract Background The oncogene epidermal growth factor receptor variant III (EGFRvIII) is expressed in approximately one third of all glioblastoma (GBM). So far it is not clear if EGFRvIII expression induces replication stress in GBM cells, which might serve as a therapeutical target. Methods Isogenetic EGFRvIII- and EGFRvIII+ cell lines with endogenous EGFRvIII expression were used. Markers of oncogenic and replication stress such as γH2AX, RPA, 53BP1, ATR and Chk1 were analysed using western blot, immunofluorescence and flow cytometry. The DNA fiber assay was performed to analyse replication, transcription was measured by incorporation of EU and genomic instability was investigated by micronuclei and CGH-Array analysis. Immunohistochemistry staining was used to detect replication stress markers and R-loops in human GBM samples. Results EGFRvIII+ cells exhibit an activated replication stress response, increased spontaneous DNA damage, elevated levels of single stranded DNA and reduced DNA replication velocity, which are all indicative characteristics for replication stress. Furthermore, we show here that EGFRvIII expression is linked to increased genomic instability. EGFRvIII expressing cells display elevated RNA synthesis and R-loop formation, which could also be confirmed in EGFRvIII positive GBM patient samples. Targeting replication stress by irinotecan resulted in increased sensitivity of EGFRvIII+ cells. Conclusion This study demonstrates that EGFRvIII expression is associated with increased replication stress, R-loop accumulation and genomic instability. This might contribute to intratumoral heterogeneity but may also be exploited for individualized therapy approaches.


2021 ◽  
Vol 15 (1) ◽  
pp. 229-235
Author(s):  
Doaa Ghorab ◽  
Ahmed Helaly ◽  
Amani E. Badawi

Introduction: Pterygium is a common ophthalmic problem in the Middle East where exposures to dust and sun rays are risk factors. The condition is more prevalent in middle-aged males and can be considered as an aging process. The aim of this study is to test both the degenerative and the proliferative components of Pterygium by both reduced glutathione and topoisomerase one activity. Methods: The study applied immunohistochemistry staining for both reduced glutathione and topoisomerase 1. Results: The samples expressed positive glutathione staining in most primary Pterygium conditions and all secondary Pterygium. On the other hand, the topoisomerase 1 immunohistochemistry expressed focal activity in secondary conditions suggesting a progenitor cell role in the pathogenesis of Pterygium in conjunction with oxidative stress. Conclusion: Pterygium represents dual pathology with a proliferative component and a degenerative one that needs further studies. It is possible to use combination immunohistochemistry markers to predict the prognosis of Pterygium behavior.


2021 ◽  
Vol 12 ◽  
pp. 560
Author(s):  
Ayu Yoniko Christi ◽  
Wisnu Baskoro ◽  
Bidari Kameswari ◽  
Irfaanstio Akbar Hakim ◽  
Vega Sola Gracia Pangaribuan ◽  
...  

Background: Malignant peripheral nerve sheath tumors (MPNSTs) typically found in the trunk, limbs, head, and neck represent 3–10% of all soft-tissue sarcomas. Although they typically originating from peripheral nerve Schwann cells, 2–3% arise from the spinal nerves and may be found within the spinal canal. Here, we present a 43-year-old male with an extradural thoracic MPNST contributing to marked cord compression and a progressive paraparesis. Case Description: A 43-year-old male presented with a progressive paraparesis of 16 months’ duration. The MRI showed a posterior T2-T4 extradural tumor in the thoracic spine resulting in significant cord compression. Following a T2-T4 laminectomy and gross total excision of the epidural mass, the patient regained modest neurological function. Immunohistochemistry staining supported the diagnosis of thoracic spinal MPNST. Conclusion: Rarely, spinal MPNST can be considered amongst the differential diagnoses of an extradural spinal tumor. In this case, gross total excision of a posterior T2-T4 epidural MPNST resulted in improvement in the patient’s original paraparesis. Notably, immunohistochemistry staining helped confirm the diagnosis of a MPNST.


2021 ◽  
Author(s):  
Nanni Hua ◽  
Chen Yang ◽  
Anxian Chen ◽  
Yi Feng ◽  
Xianglei He ◽  
...  

Abstract Background ConclusionsFibrinogen-like protein-1 (FGL1), as the member of FREP superfamily, had been widely concerned as a major immune inhibitory ligand of LAG-3. Although FGL1 expression levels didn’t have significant difference in most of tumors via online data analysis, we found that it was down-regulated in liver cancer. Moreover, the correlation between FGL1 expression and the progression and prognosis of hepatocellular carcinoma (HCC) was still disputed. Methods In our study, firstly, we used bioinformatics analysis to define the expression profile and clinical significance of FGL1 in HCC. Then, we determined the FGL1 level in human HCC cell lines, tumor and normal liver tissues from HCC patients by western blot. Furthermore, tissue microassays were used to detect the expression of FGL1 through immunohistochemistry staining and verify whether FGL1 expression levels were associated with clinicopathological features of HCC patients. Results The results proved that FGL1 was down-regulated significantly in HCC cell lines and HCC tissues, corresponding with the results of our bioinformatics analysis. FGL1 expression levels in HCC were related to Edmondson grade and metastasis. Additionally, high FGL1 expression was related to better overall survival in HCC patients, indicating that the down-regulated FGL1 was correlated with poor prognosis and FGL1 might function as a tumor suppressor. Conclusions Taken together, expression levels of FGL1 may correlate with the progression and prognosis of HCC, and FGL1 could be a potential prognostic biomarker.


2021 ◽  
Vol 13 (1) ◽  
pp. e2021063
Author(s):  
PIMJAI NIPARUCK

Background: Double-expressor lymphoma (DEL) was found accounting for 20- 30% of DLBCL. We conducted this study to analyze the survival, the clinical presentation and the factors associated with treatment outcomes in DEL-DLBCL. Methods:  A retrospective study of 291 patients diagnosed with DLBCL during January 2015 - December 2018. Results: Of the 291 patients, median age was 63 years, germinal center B cell like DLBCL (GCB) and non-GCB subtypes were found in 32% and 68%, respectively. DEL was found in 46% of 264 patients with available immunohistochemistry staining for MYC protein. Patients with DEL was significantly more common in elderly patients (p= 0.017) and non-GCB subtype (p= 0.006). High serum lactate dehydrogenase (LDH) levels and high Ki-67 index were significantly found in DEL patients than those in non-DEL patients (p= 0.024 and p= 0.04, respectively).  3y-OS and 3y-DFS in DEL patients were shorter than those in non-DEL group, 58.7% versus 78.9% (p=0.026) and 58.4% versus 67.7% (p=0.343), respectively. Independent factors affecting both OS and DFS in DEL patients were ECOG 3-4, high LDH level, extranodal involvement> 1 site, DHL, high IPI and stage III-IV. Conclusions: High incidence of DEL was observed in this study, especially in patients aged 60 years or older and non-GCB subtype. Patients with DEL showed dismal DFS and OS.


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