[Regulation of colony-stimulating factor 1 receptor inhibitor pexidartinib on the senescence of mouse bone marrow-derived macrophages stimulated by lipopolysaccharide].

Objective: To investigate the effects of colony-stimulating factor 1 receptor (CSF-1R) inhibitor pexidartinib (PLX3397) on the senescence of bone marrow-derived macrophages (BMDM) stimulated by lipopolysaccharide (LPS). Methods: BMDM were isolated and cultured from femurs and tibiae of 10 male C57BL/6 mice aged 6-8 weeks (obtained from Laboratory Animal Center of Guizhou Medical University). They were divided into blank control group, LPS group (treated with 1 µg/ml LPS for 24 h) as well as low, medium and high concentration PLX3397 pretreatment groups (treated with 100, 500 and 1 000 nmol/L PLX3397 for 4 h respectively followed by 1 µg/ml LPS for 24 h). The corresponding markers of macrophages were detected by flow cytometry. Cell viability was detected by cell counting kit-8 and cellular senescence was detected by senescence-associated-ß-galactosidase (SA-ß-gal) staining. Meanwhile, protein expressions of cycle-dependent kinase inhibitor p16, p21 and CSF-1R were detected by Western blotting, and the expressions of p16 and p21 were detected by intracellular immunofluorescence. Real-time fluorescence quantitative PCR (RT-qPCR) was used to investigate the mRNA levels of senescence-associated secretory phenotype (SASP) genes including interleukin (IL), IL-1ß, chemokine-1/10 (CXCL-1/10), matrix metalloproteinase-8 (MMP-8), and transforming growth factor-ß (TGF-ß). Results: The rate of SA-ß-gal positive staining in medium and high concentration PLX3397 pretreatment groups [(39.33±4.93)% and (36.33±3.06)% respectively] were significantly downregulated compared with LPS group [(52.00±3.00)%] (P=0.020, P=0.005). The expression of CSF-1R protein in low, medium and high concentration PLX3397 pretreatment groups were (0.74±0.18, 0.61±0.07, 0.54±0.06), all of which were significantly lower than that in LPS group (1.16±0.08) (P=0.013, P=0.002, P<0.001). The expression levels of CSF-1R mRNA in low, medium and high concentration PLX3397 pretreatment groups (1.04±0.06, 0.90±0.05, 1.18±0.08) showed similar trend (2.90±0.25) (P<0.001). The average fluorescence intensity of p16 in all PLX3397 pretreatment groups were 49.76±3.65, 48.21±1.72, 47.99±1.26 respectively, which were significantly lower than that in LPS group (66.88±5.85) (P=0.001, P<0.001, P<0.001). The average fluorescence intensity of p21 in medium and high concentration PLX3397 pretreatment groups were (34.43±3.62, 30.13±0.86), significantly lower than that in LPS group (46.82±5.33) (P=0.043, P=0.007). The expression of p16 protein in low, medium and high concentration PLX3397 pretreatment groups (0.56±0.04, 0.55±0.04, 0.35±0.19) were significantly lower than that in LPS group (0.98±0.10) (P=0.003, P=0.002, P<0.001), as well the expression of p21 protein (0.69±0.20, 0.42±0.08, 0.26±0.14) (P=0.032, P=0.002, P<0.001). According to the results of RT-qPCR, the expressions of IL-6, IL-1ß, CXCL-1, CXCL-10 and MMP-8 in PLX3397 pretreatment groups were significantly lower than those in LPS group (P<0.001), while the expression of TGF-ß increased (P<0.001). Conclusions: LPS could induce the cell senescence, increase the secretion of SASP and aggravate local inflammation by activating the CSF-1R on the cell surface of bone marrow-derived macrophages. CSF-1R inhibitor PLX3397 might attenuate CSF-1R activation associated with LPS and inhibit the senescence of bone marrow-derived macrophages induced by LPS.

[Pathogen distribution and antimicrobial resistance among lower respiratory tract infections in patients with hematological malignancies].

Objective: To investigate the pathogen distribution and antimicrobial resistance among lower respiratory tract infections in patients with hematological malignancies. Methods: Sputum samples were collected from 967 patients with hematological malignancies and lower respiratory tract infections in Department of Hematology,the Second Hospital of Shanxi Medical University from January 2017 to July 2020. The pathogens and drug sensitivity reports were carried out by automatic bacterial identification instruments. WHONET 5.6 and SPSS 20.0 softwares were used for statistical analysis. Results: A total of 961 strains of pathogens were isolated, 516 (53.7%) pathogens were Gram-negative bacteria, mainly 118 strains of Klebsiella pneumonia (12.3%), 68 strains of Pseudomonas aeruginosa (7.1%), 67 strains of Acinetobacter baumannii (7.0%),52 strains of Stenotrophomonas maltophilia (5.4%), 43 strains of Escherichia coli (4.5%), and 42 strains of Enterbacter cloacae (4.4%). There were 171 (17.8%) strains of Gram-positive bacteria and 274 (28.5%) fungi. The drug resistance rates of Pseudomonas aeruginosa and Acinetobacter baumannii to carbapenem were 22.1%-31.3%. Stenotrophomonas maltophilia was sensitive to levofloxacin, compound sulfamethoxazole and minocycline. The antimicrobial resistance rates of these three enterobacteria to carbapenems, cefoperazone/sulbactam, piperacillin/tazobactam were low (<10%). The resistant Gram-positive bacteria to ticoplanin, vancomycin and linazolamide were not detected. Conclusion: The major pathogens related to lower respiratory tract infections in patients with hematological malignancies are gram-negative bacteria in our centre. Different pathogens appear different characteristics of antimicrobial resistance.

Long-term survival case of a non-small cell lung cancer bone metastasis patient treated with bone cement, radiation and gefitinib.

OBJECTIVE: We explore the treatment of bone metastases in advanced non-small cell lung cancer (NSCLC). MATERIALS AND METHODS: We reported a 76-year-old female patient, who was diagnosed with NSCLC with bone metastasis eight years ago (stage IVA). Due to unbearable diarrhea, she refused chemotherapy, and we adopted local treatment, including local radiotherapy 50 Gy and bone cement to lumbar spinal metastases, 62 Gy local radiotherapy of primary lung tumor, TKI inhibitor gefitinib and zoledronic acid. RESULTS: She survived more than eight years and is still in follow-up. CONCLUSIONS: The median survival time for NSCLC patients with bone metastases is often less than 1 year. We reported the patient with more than eight years of survival, showed that some special cases can adopt the methods of local treatment including bone cement, treatment benefit patients, radiation therapy and targeted therapy in clinic to expand the survival.

Progress of radiotherapy after breast-conserving surgery combined with silicone prosthesis reconstruction.

Breast cancer is the malignant tumor with the highest incidence in women. The standard treatment for early breast cancer is radical surgery combined with radiotherapy, but many studies have shown that adjuvant radiotherapy after breast-conserving surgery combined with silicone prosthesis reconstruction is gradually expected to become the new standard treatment because this method can obtain a good local tumor control rate, and has a cosmetic effect. Compared with myocutaneous flap reconstruction, silicone prosthesis implantation has the advantages of less trauma, simple operation, beautiful appearance, and no overlap of donor areas during reconstruction. It is a safe and feasible surgical method without worrying about necrosis and atrophy of myocutaneous flap. This emerging combination therapy may become the best mode of early breast cancer treatment.

Comparison of IMRT and VMAT radiotherapy planning for Graves' ophthalmopathy based on dosimetric parameters analysis.

OBJECTIVE: To compare volumetric modulated arc therapy (VMAT) and intensity-modulated radiation therapy (IMRT) for the treatment of Graves' ophthalmopathy (GO) based on the dosimetric data. PATIENTS AND METHODS: 19 patients diagnosed with GO were recruited in this study. For each patient, a dual-partial-arc VMAT plans and a 7-fixed-field IMRT plans were replanned. Dosimetric parameters of the targets and organs at risk (OARs) originated from the two kinds of plans were compared and analyzed. RESULTS: Homogeneity index (HI) was superior in IMRT plans compared with VMAT (p=0.0014) but there was no significant statistical difference in conformity index (CI) between them (p=0.0673). IMRT plans revealed advantage in the OARs protection especially for the left and right lenses, optic nerves and eyeballs (p<0.05). CONCLUSIONS: VMAT and IMRT are both feasible techniques in radiotherapy in GO from the perspective of dosimetric parameters. Homogeneity and OAR protection were slightly superior in IMRT plans compared with VMAT plans.

H3K27M mutant diffuse midline glioma: a case report.

OBJECTIVE: Diffuse midline glioma with H3K27M mutation is a new tumor type of WHO central nervous system tumor classification. It often occurs in the midline structure and usually has a poor prognosis. CASE REPORT: A 38-year-old male patient presented with 2 years history of right limb with facial numbness, tumors in the left thalamic region and lateral ventricle was detected by imaging. The patient underwent the first surgery. RESULTS: The pathological examination results: Glioblastoma. He recovered well after surgery and received a total of 30 times of radiotherapy and temozolomide for one year. Fourteen months later, tumours were observed in the left thalamic region and left parieto-occipital lobe, the patient underwent the second operation. Immunohistochemistry showed: H3K27M(+). He experienced limitation of right limb movement after surgery and started taking oral apatinib 250 mg qd. After one-year, multiple tumors were found in the left brainstem, bilateral ventricles, bilateral basal ganglia, etc. The patient was given radiotherapy 7 times and then took apatinib 250 mg qd. Now the patient is still alive. CONCLUSIONS: H3K27M mutant diffuse midline glioma is characterized by diffuse infiltrative growth. Its pathological classification is diverse, imaging features lack specificity, and prognostic factors are complex. Traditional radiochemotherapy has limited effects, molecular targeted therapy, especially intervention of epigenetic regulation is being explored.

Treatability of organic fractions derived from secondary effluent by reverse osmosis membrane.

Dissolved organic matters (DOMs) from two batches of secondary effluent collected from a local water reclamation plant were fractionated using column chromatographic method with non-ionic resins XAD-8, AG MP-50 and IRA-96. Seven isolated fractions were obtained from the fractionation study and these fractions were quantified using DOC, UV(254) and SUVA values. The fractionation study revealed that the secondary effluent samples comprised about 47.3-60.6% of hydrophobic and 39.4-52.7% of hydrophilic solutes. The treatability of each isolated fraction was investigated by subjecting each fraction to reverse osmosis (RO) treatment individually. It was noted that RO process could achieve high DOC rejections for acid and neutral fractions (ranging from 80% to 98% removal) probably due to the negative charge of RO membrane. The results obtained also indicated that hydrophobicity of DOMs is significant in determining treatability of organic species by RO process. The performance of RO in terms of DOC rejection of un-fractionated secondary effluent was also investigated to assess possible effects of interactions among organic fractions on their treatability by RO process. It was noted that DOC rejection associated with the un-fractionated secondary effluent was generally higher (ranging from 2% to 45%) than the corresponding rejection obtained from each individual fraction isolated from the secondary effluent. This finding suggested there is a beneficial interaction among the fractions that in turn has contributed towards a better overall DOC rejection performance by RO treatment.

Insulin regulates protein kinase CbetaII expression through enhanced exon inclusion in L6 skeletal muscle cells. A novel mechanism of insulin- and insulin-like growth factor-i-induced 5' splice site selection.

The protein kinase Cbeta (PKCbeta) gene encodes two isoforms, PKCbetaI and PKCbetaII, as a result of alternative splicing. The unique mechanism that underlies insulin-induced alternative splicing of PKCbeta pre-mRNA was examined in L6 myotubes. Mature PKCbetaII mRNA and protein rapidly increased >3-fold following acute insulin treatment, while PKCbetaI mRNA and protein levels remained unchanged. Mature PKCbetaII mRNA resulted from inclusion of the PKCbetaII-specific exon rather than from selection of an alternative polyadenylation site. Increased PKCbetaII expression was also not likely accounted for by transcriptional activation of the gene or increased stabilization of the PKCbetaII mRNA, and suggest that PKCbetaII expression is regulated primarily at the level of alternative splicing. Insulin effects on exon inclusion were observed as early as 15 min after insulin treatment; by 20 min, a new 5'-splice site variant of PKCbetaII was also observed. After 30 min, the longer 5'-splice site variant became the predominate species through activation of a downstream 5' splice site. Similar results were obtained using IGF-I. Although the role of this new PKCbetaII mRNA species is presently unknown, inclusion of either PKCbetaII-specific exon results in the same PKCbetaII protein.

15-Hydroxyeicosatetraenoic acid dehydrogenase activity in microsomal fraction of mouse liver homogenate.

The microsomal fraction of mouse liver homogenate showed NAD(P)-dependent dehydrogenase activity involved in the conversion of 15-hydroxyeicosatetraenoic acid to 15-ketoeicosatetraenoic acid, which was determined quantitatively by HPLC assay. This enzyme, tightly bound to membranes and relatively stable, possessed apparent values of Km of 8.3 microM and Vmax of 2.8 nmoles/mg.min in the oxidation of 15-HETE, and gave an optimum pH of 9.8. Additionally, the enzyme, not susceptible to the inhibition by indomethacin and showing a similar cosubstrate specificity between NAD and NADP, utilized other hydroxylated eicosanoids as substrates, based on HPLC analyses.

Soybean lipoxygenase-catalyzed formation of lipoxin A and lipoxin B isomers from arachidonic acid via 5,15-dihydroperoxyeicosatetraenoic acid.

Soybean lipoxygenase converted arachidonic acid to a group of polar products (lambda max, 300-301 nm), which were increasingly formed during the continued incubation at 20 degrees C after the initial incubation (2 hrs, at 4 degrees C). These products were identified as lipoxin A and B isomers, based on the chromatographic and spectrometric analyses. In further chromatographic analyses, the lipoxin A and B isomers were separated into at least three isomers, respectively. The exposure of 5,15-dihydroperoxyeicosatetraenoic acid to the soybean lipoxygenase produced the identical product profile of chromatography, substantiating the intermediacy of 5,15-dihydroperoxyeicosatetraenoic acid in the soybean lipoxygenase-catalyzed formation of lipoxins. Based on these results, it is proposed that the conversion of arachidonic acid into lipoxins by soybean lipoxygenase may bear a mechanistic resemblance to the formation of lipoxins in the human leukocytes.

Use of adenine nucleotide derivatives to assess the potential of exo-active-site-directed reagents as species- or isozyme-specific enzyme inactivators. 3. Synthesis of adenosine 5'-triphosphate derivatives with N6- or 8-substituents bearing iodoacetyl groups.

Several series of N6- or 8-substituted derivatives of adenosine 5'-triphosphate (ATP) were synthesized. N6-(omega-Aminoalkyl) derivatives of adenosine 5'-monophosphate (AMP) were converted into their omega-N-carbobenzyloxy derivatives, and these were converted, via the 2',3'-O-carbonyl derivatives of their 5'-phosphorimidazolidates, into the corresponding ATP derivatives. Hydrogenolytic removal of the carbobenzyloxy groups, followed by iodoacetylation of the omega-amino groups with N-(iodoacetoxy)succinimide, gave N6-R-ATP, where R = (CH2)nNHCOCH2I (n = 2--8) or (CH2)nCON)CH3)(CH2)mN(CH3)CO(CH2)nNHCOCH2I (n = m = 3; n = 3, m = 4; n = 4, m = 3; n = m = 4). Condensation of N6-(omega-aminoalkyl) derivatives of AMP with N-hydroxysuccinimide esters of omega-[N-(carbobenzyloxy)amino] carboxylic acids gave N6-(CH2)nNHCO(CH2)mNH-Cbz derivatives of AMP which, upon conversion to the corresponding derivatives of ATP, followed by removal of the carbobenzyloxy group and iodoacetylation, as described above, gave N6-(CH2)nNHCO(CH2)mNHCOCH2I-ATP derivatives (n = 3, m = 5 or 6; n = 4, m = 5; n = 6, m = 1--6). The same sequence of reactions starting with N6-[omega-(methylamino)alkyl] derivatives of N6-CH3-AMP gave N6-CH3, N6-(CH2)nH(CH3)CO(CH2)mNHCOCH2I derivatives of ATP (n = 4, m = 3, 5 or 6; n = 6, m = 5 or 6). Reaction of alpha, omega-diaminoalkanes with 8-Br-ATP gave 8-NH(CH2)nNH2 derivatives of ATP, which upon iodoacetylation gave 8-NH(CH2)nNHCOCH2I derivatives of ATP (n = 2, 4, 6, or 8). Substrate and inhibitor properties indicated that the ATP derivatives are potential exco-ATP-site-directed inactivators of hexokinases, adenylate kinases, and pyruvate kinases.