[Role of CT and MRI image fusion and computer assisted simulation technique in guiding type â ¢ and â £ primary pelvic sarcoma surgeries].

Objective: To explore the possible clinical benefits of CT/MRI image fusion and computer-assisted simulation techniques in guiding type Ⅲ and Ⅳ primary pelvic bone tumor surgeries. Methods: The clinic data of primary bone sarcomas patients treated at Department of Bone and Soft Tissue,Zhenghzhou University Affiliated Cancer Hospital from January 2019 to December 2021 were retrospectively analyzed. Based on whether the CT and MRI image fusion technique was utilized for tumor evaluation and surgical planning,the patients were divided into image fusion group (n=21) or control group (n=27). There were 7 male and 14 female patients included in the image fusion group, with the age of (37.0±10.4) years(range: 18 to 67 years). In the control group, there were 10 males and 17 females with the age of (39.7±15.2) years (range: 16 to 65 years). Both groups included osteosarcoma,chondrosarcoma and undifferentiated polymorphic sarcoma as the pathological diagnosis. Clinical information such as gender,age,pathological diagnosis,location of disease,and metastasis at diagnosis were collected. Surgical related information such as duration of surgery,blood loss,surgical margin,and wound complications were also obtained. Periodical follow-ups every 3 months were performed for all patients to monitor the status of local recurrence,distant metastasis,and survival information. Independent t test and χ² test were used for data comparison between groups. Results: Significant reduced duration of surgery was observed in the image fusion group in comparison with control group both in type Ⅲ and Ⅳ surgeries ((144.0±31.6)min vs. (248.2±56) min,t=-8.084, P<0.01); (173.0±42.0)min vs. (306.1±62.0)min, t=-4.518, P<0.01). Blood loss was significantly reduced in the image fusion group compared with the control group ((484.8±226.3)ml vs. (836.1±359.8)ml,t=-4.130, P<0.01). In addition, significant lower ratio of R1 margin and recurrence rates of type Ⅲ and Ⅳ surgeries were found in the image fusion group comparing with the control group (4.8%(1/21) vs. 22.2%(6/27), χ²=4.214, P=0.040; 4.8%(1/21) vs. 22.2%(6/27), χ²=4.214, P=0.040).In the image fusion group, there were 3 cases of incision infection, 1 of which underwent secondary debridement.And in thecontrol group there were 7 cases of incision infection, 3 of which underwent secondary debridement. There was no significant difference in the incidence of complications between the two groups (14.2%(3/21)vs. 25.9%(7/27), χ²=0.645, P=0.422). Up to the last follow-up, 1 patient died in the image fusion group and 2 patients died in the control group, the difference was not statistically significant (χ²=1.885, P=0.220). Conclusion: Compared with the traditional operation,the image fusion technique can significantly reduce the duration of surgery,blood loss and lower the recurrence rate by achieving better surgical margins.

[Research progress of micronucleus visualization analysis and artificial intelligence detection strategy].

The micronucleomics test can comprehensively display a variety of harmful endpoints, such as DNA damage and repair, chromosome breakage or loss and cell growth inhibition, with fast, simple and economical feature. Micronucleomics is not only widely used in the comprehensive assessment of the types and modes of genetic action of exogenous chemicals (such as drugs, food additives, cosmetics, environmental pollutants, etc.), but also plays an important role in the screening and risk assessment of cancer population at high risk. However, the traditional micronucleomics image counting method has the characteristics of time-consuming, low accuracy, and high cost, which cannot meet the current analysis requirements of large-scale, multi-index, rapidity, high precision and visualization. In recent years, with the rapid development of the era of precision medicine based on big data, visualized analysis of new micronucleomics based on machine learning and detection strategies based on deep learning have shown a good application prospect. This review, based on the application value of micronucleomics, systematically compares the traditional and new artificial intelligence counting of micronucleus images, and discusses the future direction of micronucleus image detection.

[Inhibition of receptor-interacting protein 3 improves experimental autoimmune hepatitis].

Objective: To evaluate the potential of receptor-interacting protein 3 (RIP3) as a therapeutic target for autoimmune hepatitis (AIH). Methods: Immunofluorescence assay was used to observe the activated expression levels of RIP3 and its downstream signal mixed lineage protein kinase domain-like protein (MLKL) in the liver tissues of patients with AIH and hepatic cyst. Concanavalin A (ConA) was injected into the tail vein to induce acute immune-mediated hepatitis in mice. Intervention was performed by intraperitoneal injection of RIP3 inhibitor GSK872 or solvent carrier. Peripheral blood and liver tissues were collected. Serum transaminases level, qPCR and flow cytometry were analyzed. The intergroup comparison was performed with an independent sample t-test. Results: The expression level of p-RIP3 (the activated forms of RIP3) and phosphorylated p-MLKL (MLKL after phosphorylation) downstream signal were significantly higher in the liver tissue of AIH patients than those of controls. Compared with the control group, the expression levels of RIP3 and MLKL mRNA were significantly increased in the liver tissue of AIH patients (relative expression levels 3.28±0.29 vs. 0.98±0.09, 4.55±0.51 vs. 1.06±0.11), and the differences were statistically significant (t=6.71 and 6.77, respectively, and P<0.01). The expression levels of RIP3 and MLKL mRNA were significantly higher in the mice liver tissue of ConA-induced immune hepatitis than those in the control group (relative expression levels 2.35±0.09 vs. 0.89±0.11,2.77±0.22 vs. 0.73±0.16,t=10.4,6.33, P<0.01). RIP3 inhibitor GSK872 had significantly attenuated ConA-induced immune liver injury and inhibited the expression of tumor necrosis factor-α, interleukin-6, interleukin-1ß and NLRP3 in liver. Compared with the control group, the proportions of CD45+F4/80+ macrophages, CD4+ IL-17+ Th17 cells, CD4+ CD25+ regulatory T (Treg) cells and CD11b+ Gr-1+ myeloid derived suppressor cells (MDSCs) were significantly increased in the liver of ConA + Vehicle group. Compared with ConA + Vehicle group, the proportion of CD45+F4/80+ macrophages and CD4+ IL-17+ Th17 cells were significantly decreased, while the proportion of CD4+ CD25+Treg cells and CD11b+ Gr-1+ MDSCs with immunomodulatory functions were significantly increased in mice liver of ConA+GSK872 group. Conclusion: AIH patients and ConA-induced immune hepatitis mice have activated RIP3 signal in liver tissues. Inhibition of RIP3 reduces the expression and proportion of proinflammatory factors and cells, and promotes the accumulation of CD4+ CD25+ Treg cells and CD11b+ Gr-1+ MDSCs with immunomodulatory functions in the liver of mice with immune hepatitis, thereby alleviating liver inflammation and injury. Therefore, the inhibition of RIP3 is expected to be a new approach for the treatment of AIH.

[Characteristics of initial diagnosis in autoimmune liver disease: an 18-year, retrospective single-center study].

Objective: To analyze the chorological changes of diagnosis in patients with autoimmune liver disease (AILD) and related factors for early diagnosis. Methods: A total of 581 patients with age ranged from 16 to 81 were retrospectively analyzed, who were admitted to Tianjin Medical University General Hospital with AILD during January 2000 to December 2017. Age at diagnosis, diagnostic method and cirrhosis at diagnosis were compared in different groups according to admission period as 2000-2005, 2006-2011, 2012-2017. Results: The diagnostic rate of AILD showed an upward trend during the past near two decades. The proportion of AILD patients diagnosed via health examination was increasing year by year mainly by elevated transaminases (P<0.001). The mean age at diagnosis in our AILD patients were younger at present, especially in men (P=0.044). The proportion of cirrhosis at diagnosis was gradually reduced in three different periods respectively [77.78%(21/27), 41.58% (79/190), 25.00%(91/364), P<0.001], which were coincident in patients with autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) (P<0.001). The shrinking trend of cirrhosis at diagnosis was significantly correlated with the increasing application of health examination (r=-0.549, P<0.001). Conclusions: Extensive application of health examination expands the diagnostic rate of AILD. During the past 18 years, more young patients are diagnosed with AILD. The proportion of severe cases such as cirrhosis at diagnosis is decreasing. Screening of immunological examinations in patients with abnormal transaminases is needed and critical to the early diagnosis of asymptomatic AILD.

MiR-485-5p inhibits metastasis and proliferation of osteosarcoma by targeting CX3CL1.

OBJECTIVE: To investigate the potential effect of miR-485-5p on the development of osteosarcoma (OA) and its relevant mechanism. PATIENTS AND METHODS: The expression level of miR-485-5p was detected in OA tissues and cells (MG-63) comparing with corresponding adjacent normal tissues and normal human osteoblastic cell lines (Hfob1.19), respectively. Luciferase assay was performed to evaluate the interaction between miR-485-5p and CX3CL1, the effects of miR-485-5p on MG-63 cells were determined by subsequent experiments including cell proliferation, expression level of CX3CL1, detection of invasion and migration capacities. RESULTS: In our present research, miR-485-5p was down-regulated in OA tissues and we got the same result in OA cells. In order to obtain potential target of miR-485-5p, we checked it in three publicly available algorithms, TargetScan, miRDB and microRNA. We found that CX3CL1 is a direct target of miR-485-5p, and Luciferase assays confirmed our hypothesis. The results showed that decreased expression of CX3CL1 resulting from the up-regulation of miR-485-5p could decelerate cell proliferation, invasion and migration in OA cells. CONCLUSIONS: We showed the suppressor function of miR-485-5p in OA by targeting CX3CL1, indicating that miR-485-5p/CX3CL1 axis might be a potential therapeutic target for the treatment of OA.

[Role of Rac1 signaling pathway of azathioprine and peptidoglycan in the regulation of monocyte-macrophage apoptosis in Crohn's disease].

Objective: To evaluate the changes of macrophages and expression of Rac1 in the inflammatory site of Crohn's disease, and to investigate the effects of 6-thioguanine (6-TG) and peptidoglycan on apoptosis of human peripheral blood monocyte-macrophage by regulating Rac1 signaling pathway. Methods: Ten patients with Crohn's disease and eight healthy controls diagnosed were enrolled at Department of Gastroenterology and Hepatology, Tianjin Medical University General Hospital from January 2013 to January 2014. The number of macrophages, apoptosis and expression of Rac1 in the inflammation sites and non-inflammation sites of intestinal mucosa were detected in both patients and controls. Peripheral blood mononuclear cells (PBMCs) were sorted by CD14 immunomagnetic beads. The apoptosis of monocytes, expression of Rac1 and related apoptosis signaling molecules were detected in patients treated with peptidoglycan, 6-TG and Rac1 inhibitor NSC23766 and another 15 healthy donors. Results: The number of macrophages and apoptotic cells significantly increased in the inflammatory group of Crohn's disease patients compared with the non-inflammatory group. The expression of PAK1, downstream molecular of Rac1 signaling pathway of macrophages was also significantly higher in the inflammatory group of Crohn's disease patients than that in healthy controls and non-inflammatory group. Compared with control group, anti-apoptotic signals (NF-κB, Bcl-xL and STAT-3) in PBMCs increased in the peptidoglycan group, while slightly decreased in 6-TG group. 6-TG and NSC23766 significantly promoted peptidoglycan-related anti-apoptosis [peptidoglycan group (8.6±3.7)%, peptidoglycan+ 6-TG group (42.0±2.7)%, peptidoglycan+ NSC23766 group (58.5±6.9)%, P<0.05]. Conclusions: Peptidoglycan plays a role in the pathogenesis of Crohn's disease by recruiting macrophages. However, 6-TG inhibits peptidoglycan-induced activation of Rac1 signaling pathway leading to macrophage apoptosis.

[The study of liver macrophages polarization in patients with autoimmune hepatitis].

To explore the functional phenotype of liver macrophages in patients with autoimmune hepatitis (AIH). Compared with patients with nonalcoholic fatty liver disease (NAFLD) and chronic hepatitis B (CHB), the absolute CD(68)(+) cell count in patients with AIH was significantly higher (all P<0.05). It was positively correlated with ALT and IgG levels (the correlation coefficients 0.600 and 0.700, P=0.285 and 0.188 respectively). Additionally, compared with patients with NAFLD, the absolute iNOS positive cell count in patients with AIH and CHB were significantly higher (all P<0.05). The expression of TNFα, iNOS and IL-1ß in patients with AIH and CHB were significantly higher than in patients with NAFLD (all P<0.05). Interestingly, compared with patients with AIH and CHB, the absolute CD(206)(+) cell count in patients with NAFLD were significantly higher (all P<0.05). CD(206) expression in patients with NAFLD was higher than patients with AIH and CHB, but with no statistical significance. M1 type macrophages over-expressed and played a major role in the inflammatory reaction and liver injury in patients with AIH.

Lactobacillus rhamnosus GG supernatant upregulates serotonin transporter expression in intestinal epithelial cells and mice intestinal tissues.

BACKGROUND: The role that probiotics play in relieving irritable bowel syndrome (IBS) has been demonstrated; however, the mechanism by which IBS is affected remains unclear. In this study, serotonin transporter (SERT) mRNA and serotonin transporter protein (SERT-P) levels in HT-29, Caco-2 cells, and mice intestinal tissues were examined after treatment with Lactobacillus rhamnosus GG supernatant (LGG-s). METHODS: HT-29 and Caco-2 cells were treated with different concentrations of LGG-s for 12 and 24 h and C57BL/6 mice received supplements of different concentrations for 4 weeks. SERT mRNA and SERT-P levels were detected by real-time PCR and Western blotting. KEY RESULTS: SERT mRNA and SERT-P levels in HT-29 and Caco-2 cells were higher than those in the control 24 h after treatment. Undiluted LGG-s upregulated SERT mRNA levels by 9.4-fold in the first week, which dropped in the second week. The double-diluted LGG-s upregulated SERT mRNA by 2.07-fold in the first week; levels dropped to 1.75-fold within the second week and under base expression levels by the third week, while they again climbed to 1.56-fold in the fourth week. The triple-diluted LGG-s could not upregulate SERT mRNA expression until the end of the fourth week. The SERT-P levels in the double-diluted LGG-s group were higher than that in the control but fluctuated with time. SERT-P levels in the triple-diluted LGG-s were higher than that in the control in the last 2 weeks and increased with time. CONCLUSIONS & INFERENCES: LGG-s can upregulate SERT mRNA and SERT-P levels in intestinal epithelial cells and mice intestinal tissues.

Adenovirus-mediated GM-CSF gene and cytosine deaminase gene transfer followed by 5-fluorocytosine administration elicit more potent antitumor response in tumor-bearing mice.

Antitumor effects of combined transfer of suicide and cytokine genes were investigated in this study. Adenovirus harboring E. coli cytosine deaminase gene (AdCD) and adenovirus harboring murine granulocyte-macrophage colony-stimulating factor gene (AdGMCSF) were used simultaneously for in vivo gene transfer in melanoma-bearing mice. Growth inhibition of established tumors and prolongation of survival period were observed more significantly in tumor-bearing mice after transfection with AdGMCSF and AdCD followed by continuous injection of prodrug 5-fluorocytosine (5FC) when compared with mice treated with control adenovirus AdlacZ/5FC, AdCD/5FC or AdGMCSF alone (P < 0.01). After combined therapy the expression of MHC-I (H-2Db) and B7-1 molecules on freshly isolated tumor cells increased greatly and more dendritic cells and CD8+ T cells infiltrated into the tumor mass. The activity of specific cytotoxic T lymphocytes was also found to be induced more significantly after the combined therapy. Further experiments showed that apoptosis of tumor cells and induction of antitumor immune response might be involved in the mechanisms of the tumor cell killing by the combined therapy. Our results demonstrated that combined transfer of the GM-CSF and CD suicide genes, being able to inhibit the growth of melanoma synergistically and induce specific antitumor immune response efficiently, thus addressing the drawbacks of suicide gene therapy or cytokine gene therapy which were proved to be not satisfactory when used alone, might be of therapeutic potential for gene therapy of cancer.

Adenovirus-mediated combined suicide gene and interleukin-2 gene therapy for the treatment of established tumor and induction of antitumor immunity.

The antitumor effect of the combined transfer of a suicide gene and a cytokine gene was evaluated in the present study. Adenoviruses expressing Escherichia coli cytosine deaminase (AdCD) and adenoviruses expressing murine interleukin-2 (AdIL-2) were utilized for the treatment of established tumors. The mice were inoculated s.c. with FBL-3 erythroleukemia cells and 3 days later received an intratumoral injection of AdCD in the presence or absence of AdIL-2 followed by intraperitoneal 5-fluorocytosine (5-FC) administration. The results demonstrated that tumor-bearing mice treated with AdCD/5-FC in combination with AdIL-2 showed more potent inhibition of tumor growth and survived much longer than did mice treated with AdCD/5-FC, AdIL-2, adenovirus expressing beta-galactosidase/5-FC or phosphate-buffered saline. The tumor mass showed obvious necrosis and inflammatory cell infiltration, and more CD4+ and CD8+ T cells infiltrating the tumor after combined therapy. The splenic natural killer and cytotoxic T lymphocyte activities increased significantly in the mice after combined therapy with AdCD/5-FC/AdIL-2. Our results demonstrate that therapy combining a suicide gene and IL-2 gene can inhibit the growth of established tumors in mice significantly and induce antitumor immunity of the host efficiently.

[Enzyme cytochemistry and morphometric study of the effects of macrophages on A549 pulmonary alveolar cell carcinoma cell line].

In order to study the mechanism of the effects of M phi on tumor cells, enzyme cytochemistry and morphometry were used to investigate the activities of cytochrome oxidase (CO), succinate dehydrogenase (SD), lactate dehydrogenase (LDH) and acid phosphatase (ACP) in A549 pulmonary alveolar cell carcinoma cells which had been interacted with normal and CP-activated macrophages respectively. It was found that when E/T = 10:1, the enzyme activity of the cancer cell mitochondria, CO, SD, LDH were decreased, and when E/T = 20:1, the activity of the lysosomal enzyme ACP was increased. These results indicate that when the E/T ratio was appropriate, activated M phi may injure the mitochondria and lysosomes and affect the aerobic respiration and oxidative phosphorylation of cancer cells. This may be one of the cytostatic and cytotoxic mechanisms of activated M phi on tumor cells.

[Effect of recombinant human stem cell factor (rhSCF) on self-renewal of stem/progenitor cells of human acute myelogenous leukemia].

Using serum-free culture system and liquid/semi-solid dual culture technique, we examined the effect of rhSCF on self-renewal of the stem/progenitor cells of human acute myelogenous leukemia (AML-CFU). The results revealed that the rhSCF as a hematopoietic growth factor (HGF) exerted its effect hierarchically earlier than interleukin-3 and granulocyte/macrophage colony-stimulating factor on AML-CFU. rhSCF, whether used alone or in combination with other HGFs, showed potential maintenance of self-renewal of AML-CFU in most AML patients studied. However, considerable discrepancy existed in maintenance among various FAB types of patients or even in patients with the same FAB type. It is suggested that SCF may play an important role in the pathological development of AML.

A newly developed serum-free culture system: clonal growth of human acute myelogeneous leukemia (AML) progenitors--a report of 14 AML cases.

A new serum-free culture (SFC) system for human AML-CFU was established and the colony-promoting activity of four recombinant human hematopoietic growth factors (rhHGFs) including granulocyte-macrophage colony-stimulating factor (rhGM-CSF), interleukin-3 (rhIL-3), erythropoietin (rhEPO) and newly developed stem cell factor (rhSCF) were investigated in this SFC system. Under the orthogonal design condition, it was found that human AML-CFU presented optimal clonal growth in an environment of bovine serum albumin (0.6%), saturated human transferrin (2 x 10(-6) mol/L), cholesterol (2.8 micrograms/ml), bovine insulin (15 micrograms/ml), bovine hemin (0.05 mmol/L), linoleic acid (2.8 micrograms/ml), and IMDM. Spontaneously growing colonies were observed in 11 out of 14 cases studied. The plating efficiencies obtained by culturing with rhGM-CSF, rhIL-3, and rhSCF were 0.776 +/- 0.621%, 0.574 +/- 0.510%, and 0.647 +/- 0.543% (mean +/- s), respectively. There was one case (M3b) showing no response to all HGFs in both SFC ad SCC. The clonal growth of AML-CFU obtained from peripheral blood of the patient with M6 was unexpectedly marked. As a whole, the newly designed SFC system has been demonstrated to be useful for culture of human AML-CFU from both bone marrow and peripheral blood.

An effective method for depleting mature T lymphocytes from bone marrow cells--two-step Percoll centrifugation.

In the present paper we have observed the effect of discontinuous gradient two-step Percoll centrifugation on depleting mature T lymphocytes from normal bone marrow. The pre-/post-Percoll percentage of CD34+, and Leu4+ cells in MNC was counted by using APAAP technique. As the two-step Percoll centrifugation is simple, and time saving, and decreases the incidence of contamination of cultured cells as well, This technique may be of value in serum-free culture of hematopoietic cells and immunological studies.

The significance of Ki-67, BrdU and OKT9 expression in determination of myelodysplastic syndromes by use of APAAP technique.

Monoclonal antibodies (McAb) to Ki-67, BrdU and OKT9 were used in APAAP technique for determining the proliferative cells of 46 myelodysplastic syndromes (MDS) with 16 serving as normal controls. According to Ki-67 positive labelling, the proliferative fractions of bone marrow cells can be divided into low (< 25%), intermediate (25%-35%), and high (> 35%) grades. The labelling positivity of Ki-67, BrdU and OKT9 in MDS was all higher than that of the controls. Furthermore, the positive percentages of these three McAbs were significantly correlated. From the results of our experiment, an increase in proliferative fraction was related to clinical classification, prognosis, and therapy of MDS. On the other hand, the positive percentage of Ki-67 > 40% and BrdU > 30% or ratio of BrdU to Ki-67 > 80% may be considered as parameters for unfavourable prognosis.