Image 1.

AIM:To investigate the role of ClC-3 chloride channel in the promotion of radio sensitization of na-sopharyngeal carcinoma CNE-2Z cells by dihydroartemisinin(DHA).METHODS:MTT was used to detect the inhibito-ry effect of DHA on the viability of CNE-2Z cells and normal nasopharyngeal epithelial NP69-SV40T cells,the radio sensi-tization effect of DHA on CNE-2Z cells was detected by cloning assay,the expression of ClC-3 protein was detected by Western blot,the expression of ClC-3 protein was down-regulated by siRNA technology,and the chlorine current of cells was recorded by whole cell patch-clamp technology.RESULTS:(1)Compared with NP69-SV40T cells,DHA selective-ly inhibited the proliferation of CNE-2Z cells,with IC10 values of(13.020±4.831)μmol/L and(5.244±1.050)μmol/L,respectively(P＜0.01).(2)The results of clonal formation experiments showed that DHA had a radio sensitizing effect on CNE-2Z cells,with a radio sensitization ratio of 1.9.(3)DHA could activate the chlorine channel of CNE-2Z cells and produce an outward chlorine current,but had no effect on the chlorine channel of NP69-SV40T cells.(4)DHA promoted the expression of ClC-3 chloric channel protein in CNE-2Z cells(P＜0.01).(5)Chlorine channel blocker NPPB could in-hibit the radio sensitizing effect of DHA on CNE-2Z cells by 1.84 times,and also inhibited the chlorine current activated by DHA.(6)the down-regulation of CNE-2Z ClC-3 protein could inhibit the radio sensitization effect of DHA on CNE-2Z cells by 4.19 times,and the activation of chlorine current by DHA on CNE-2Z cells was no longer produced.CONCLU-SION:DHA has a radio sensitizing effect on nasopharyngeal carcinoma CNE-2Z cells,which is likely to be related to the activation of ClC-3 chloride channel.

Polymyxin B and polymyxin E (colistin) are presently considered the last line of defense against human infections caused by multidrug-resistant Gram-negative organisms such as carbapenemase-producer Enterobacterales, Acinetobacter baumannii, and Klebsiella pneumoniae. Yet resistance to this last-line drugs is a major public health threat and is rapidly increasing. Polymyxin S₂ (S₂) is a polymyxin B analogue previously synthesized in our institute with obviously high antibacterial activity and lower toxicity than polymyxin B and colistin. To predict the possible resistant mechanism of S₂ for wide clinical application, we experimentally induced bacterial resistant mutants and studied the preliminary resistance mechanisms. Mut-S, a resistant mutant of K. pneumoniae ATCC BAA-2146 (Kpn2146) induced by S₂, was analyzed by whole genome sequencing, transcriptomics, mass spectrometry and complementation experiment. Surprisingly, large-scale genomic inversion (LSGI) of approximately 1.1 Mbp in the chromosome caused by IS26 mediated intramolecular transposition was found in Mut-S, which led to mgrB truncation, lipid A modification and hence S₂ resistance. The resistance can be complemented by plasmid carrying intact mgrB. The same mechanism was also found in polymyxin B and colistin induced drug-resistant mutants of Kpn2146 (Mut-B and Mut-E, respectively). This is the first report of polymyxin resistance caused by IS26 intramolecular transposition mediated mgrB truncation in chromosome in K. pneumoniae. The findings broaden our scope of knowledge for polymyxin resistance and enriched our understanding of how bacteria can manage to survive in the presence of antibiotics.

Objective:To provide references for disease diagnosis, treatment and aeromedical assessment by analyzing the characteristics of neurosurgical diseases in hospitalized military flying personnel.Methods:The case data and aeromedical assessment conclusions of 56 military flying personnel admitted to the Neurosurgery Department of Air Force Medical Center from 2010 to 2020 were collected. The diagnosis and treatment, as well as the flight post and aircraft type were retrospectively analyzed against the assessment conclusions.Results:The constituent ratio descending order of the neurosurgical diseases in the flying personnel was cerebrovascular disease (35.71%), central nervous system tumors (17.86%), sellar lesions (17.86%), arachnoid cysts (16.07%), traumatic brain injury (5.36%), scalp tumors (3.57%) and syringomyelia (3.57%) respectively. Among the 20 patients with cerebrovascular diseases, 5 received surgical treatment and other 15 received conservative treatment. In which 10 cases were qualified or restricted qualified, 9 cases were temporarily grounded, and 1 case was disqualified for flight. Among the 10 cases with central nervous system tumor, 5 received surgical treatment, 1 received radiotherapy and 4 received conservative treatment. In which 3 cases were qualified or restricted qualified, 3 cases were temporarily grounded and 4 cases were disqualified for flight. Among the 10 cases with saddle area lesions, 2 received surgical treatment, 3 received medical treatment and other 5 were observed by followed up. In which 8 cases were qualified or restricted qualified, 1 was temporarily grounded and 1 was disqualified for flight. Among the 9 cases with arachnoid cysts, 1 received surgical treatment and 8 received conservative treatment. In which 5 cases were qualified or restricted qualified, 1 was temporarily grounded and 3 were disqualified for flight. Among the other 7 cases, 5 received medical treatment or observed by follow-up, and 2 received surgical treatment. In which 6 cases were qualified for flight and 1 was temporarily grounded.Conclusions:Neurosurgical diseases should be paid more for flight safety due to quite a few restricted qualified, temperedly grounding or disqualified cases existed in the military flying personnel with such diseases.

Objective:To provide references for disease diagnosis, treatment and aeromedical assessment by analyzing the characteristics of neurosurgical diseases in hospitalized military flying personnel.Methods:The case data and aeromedical assessment conclusions of 56 military flying personnel admitted to the Neurosurgery Department of Air Force Medical Center from 2010 to 2020 were collected. The diagnosis and treatment, as well as the flight post and aircraft type were retrospectively analyzed against the assessment conclusions.Results:The constituent ratio descending order of the neurosurgical diseases in the flying personnel was cerebrovascular disease (35.71%), central nervous system tumors (17.86%), sellar lesions (17.86%), arachnoid cysts (16.07%), traumatic brain injury (5.36%), scalp tumors (3.57%) and syringomyelia (3.57%) respectively. Among the 20 patients with cerebrovascular diseases, 5 received surgical treatment and other 15 received conservative treatment. In which 10 cases were qualified or restricted qualified, 9 cases were temporarily grounded, and 1 case was disqualified for flight. Among the 10 cases with central nervous system tumor, 5 received surgical treatment, 1 received radiotherapy and 4 received conservative treatment. In which 3 cases were qualified or restricted qualified, 3 cases were temporarily grounded and 4 cases were disqualified for flight. Among the 10 cases with saddle area lesions, 2 received surgical treatment, 3 received medical treatment and other 5 were observed by followed up. In which 8 cases were qualified or restricted qualified, 1 was temporarily grounded and 1 was disqualified for flight. Among the 9 cases with arachnoid cysts, 1 received surgical treatment and 8 received conservative treatment. In which 5 cases were qualified or restricted qualified, 1 was temporarily grounded and 3 were disqualified for flight. Among the other 7 cases, 5 received medical treatment or observed by follow-up, and 2 received surgical treatment. In which 6 cases were qualified for flight and 1 was temporarily grounded.Conclusions:Neurosurgical diseases should be paid more for flight safety due to quite a few restricted qualified, temperedly grounding or disqualified cases existed in the military flying personnel with such diseases.

Objective To explore the variation trend of natural killer (NK) cell subsets in the recipients infected with cytomegalovirus (CMV) after renal transplantation. Methods Clinical data of 92 renal transplant recipients were retrospectively analyzed. All recipients were divided into the CMV infection group (n=43), CMV infection recovery group (n=13), stable renal function group (n=15), rejection group (n=11) and other infection group (n=10). In addition, healthy adult volunteers were enrolled in the healthy control group (n=15). The proportion of NK cells in peripheral blood, the expression proportion and the mean fluorescence intensity (MFI) of CD226 and CD16 in NK cells were observed and statistically compared among different groups. Results The proportion of NK cells was 4.9% (2.2%, 11.5%) in the CMV infection group and 3.7% (2.3%, 6.5%) in the CMV infection recovery group, which were significantly lower than those in the other groups (all P < 0.05). The expression proportion of CD226 and CD16 in NK cells in the CMV infection group was significantly lower compared with those in the healthy control group and stable renal function group(all P < 0.05). The expression proportion of CD226 and CD16 in NK cells in the CMV infection recovery group was remarkably higher than those in the CMV infection group (both P < 0.05). The MFI of CD226 and CD16 in the CMV infection group was significantly lower than those in the healthy control group (both P < 0.05). The MFI of CD226 and CD16 in the CMV infection recovery group was significantly higher than those in the CMV infection group (both P < 0.05). Conclusions The expression proportion and MFI of CD226 and CD16 in NK cells are down-regulated in CMV infection period, whereas up-regulated during the CMV infection recovery period, prompting that CD226 and CD16 expressed by NK cells are intimately correlated with the course of CMV infection.

Objective To investigate the effect and mechanism of interleukin (IL)-17C in mice undergoing kidney transplantation. Methods The life-supporting kidney transplantation mice models were established using Balb/c (H-2K^d) mice as the donors, IL-17C gene knock out (IL-17CKO) mice (knockout group) and C57BL/6J(H-2K^b) mice (wild group) were chosen as the recipients. The postoperative body mass and survival time of mice were statistically compared between two groups. Pathological examination of the kidney graft was performed by using hematoxylin-eosin (HE) staining and periodic acid-Schiff (PAS) staining. The expression levels of granzyme B, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, IL-6 and IL-1β messenger ribonucleic acid (mRNA) in the kidney graft tissue were quantitatively measured by reverse transcription polymerase chain reaction (RT-PCR). The proportion of inflammatory cell infiltration in the kidney graft tissue was detected by flow cytometry. Results In the knockout group, the survival time of mice after kidney transplantation was significantly shorter than that of the wild mice (P=0.031). The body mass was more evidently decreased in the knockout group with no statistical significance from that in the wild group. Pathological examination demonstrated that the kidney graft injury in the knockout group was significantly worse than that in the wild group. The mRNA expression levels of granzyme B, IFN-γ, TNF-α, IL-6 mRNA in the knockout group were significantly up-regulated compared with those in the wild group (all P < 0.01). The mRNA expression level of IL-1β showed a decreasing trend with no statistical significance (P=0.16). Flow cytometry analysis revealed that the infiltration of CD45⁺CD11b⁺Ly6G⁺ neutrophil and CD45⁺CD11b⁺Ly6C^hi monocyte in the kidney graft of knockout mice was significantly higher compared with that of the wild mice (P < 0.05, P < 0.01), whereas the infiltration of CD45⁺Ly6C^hiF4/80⁺ macrophage did not significantly differ between two groups (P > 0.05). Conclusions IL-17C participates in the regulation of inflammatory response after kidney transplantation. It can alleviate acute rejection and improve the survival of kidney graft by down-regulating the expression of pro-inflammatory cytokines and infiltration of inflammatory cells.

As d-amino acids play important roles in the physiological metabolism of bacteria, combination of d-amino acids with antibiotics may provide synergistic antibacterial activity. The aim of the study was to evaluate and activity of d-serine alone and in combination with -lactams against methicillin-resistant (MRSA) strains, and to explore the possible sensitization mechanisms. The activity of d-serine, -lactams alone and in combinations was evaluated both by standard MICs, time-kill curves and checkerboard assays, and by murine systemic infection model as well as neutropenic thigh infection model. An synergistic effect was demonstrated with the combination of d-serine and -lactams against MRSA standard and clinical strains. Importantly, the combinations enhanced the therapeutic efficacy in the animal models as compared to -lactam alone groups. Initial mechanism study suggested possible revision of d-alanine-d-alanine residue to d-alanine-d-serine in peptidoglycan by adding of d-alanine in the medium, which may cause decreased affinity to PBPs during transpeptidation. In conclusion, d-serine had synergistic activity in combination with -lactams against MRSA strains both and . Considering the relatively good safety of d-serine alone or in combination with -lactams, d-serine is worth following up as new anti-MRSA infection strategies.

(E)-Methyl-4-aryl-4-oxabut-2-enoate (YH-8) is a novel PKnB protein kinase inhibitor with good anti-tuberculosis activity. To evaluate its pharmacokinetics in rats, a sensitive and selective high performance liquid chromatography-tandem mass spectrometric (LC--MS/MS) method has been developed and validated for the quantification of YH-8 in rat plasma for the first time. Samples were pre-treated using a liquid--liquid extraction with ethyl acetate and the chromatographic separation was performed on a C18 column by gradient elution with methanol--water as the mobile phase. YH-8 was detected using a tandem mass spectrometer in positive selected reaction monitoring (SRM) mode. Method validation revealed good linearity over the range of 1-500 ng/mL for YH-8 with a lower limit of quantification (LLOQ) of 1 ng/mL. Intra- and inter-day precision of YH-8 assay in rat plasma samples were 2.0%-6.8%, with accuracy of the method being 100.69%-106.18%. Stability test showed that when spiked into rat plasma, YH-8 was stable for 12 h at room temperature, for up to 15 days at -70 °C, and after three freeze-thaw cycles. Extracted samples were found to be stable over 12 h in an auto-sampler. The method was successfully applied to the pharmacokinetic study of YH-8 in rats after oral administration at 100 mg/kg and 200 mg/kg.

Objective To investigate the phenotypic and genetic characteristics of the lysostaphin‐resistant Staphylococcus aureus variants induced by recombinant lysostaphin in vitro .Methods Three clinical isolates of S . aureus ,including two resistant to methicillin (MRSA ) and one susceptible to methicillin (MSSA ) were induced by treatment with sub‐MIC of recombinant lysostaphin via one‐step selection in vitro .Susceptibility of the variants to antibiotics were determined and compared with their parental strains .The full length of femABX genes was amplified by polymerase chain reaction and sequenced to identify the potential mutation sites in these genes .The growth‐curve in liquid medium and virulence in a mouse systemic infection model of both parental and variant strains were observed . Results The frequency of lysostaphin resistance in S . aureus was between 10-4 to 10-8 following induction by lysostaphin . Resistance to lysostaphin was associated with a significant decrease in growth rate in vitro and virulence in vivo ,as well as increased susceptibility toβ‐lactams evidenced by the M IC of β‐lactams against the variants as low as 1/4 000 to 1/2 of the M IC against their parental strains . Sequencing of f emA BX genes showed mutation in femA gene in both variants ,which resulted in a premature termination codon .Conclusions Resistance of S . aureus to lysostaphin may develop following induction by recombinant lysostaphin in vitro . The lysostaphin‐resistant S . aureus variants are characteristic of lower growth rate , decreased virulence ,and higher susceptibility to β‐lactams .