ObjectiveTo investigate the reversal effect of Cordyceps sinensis extract on immunotherapy resistance of non-small cell lung cancer （NSCLC）. MethodsELISA， flow cytometry， and co-culture assays were performed to evaluate the effects of Cordyceps sinensis extract on cytokine secretion， proliferation， and tumor cell killing activity of Jurkat （JKT） cells. Furthermore， the therapeutic effect of Cordyceps sinensis extract on programmed cell death ligand 1 （PD-L1） resistant lung cancer was evaluated through in vivo experiments both alone and in combination with anti-PD-L1 monoclonal antibody. ResultsCordyceps sinensis extract enhanced the secretion of cytokines interferon gamma（IFN-γ）， tumor necrosis factor-alpha（TNF-α） and interleukin-2 （IL-2）in JKT cells， promoted cell proliferation （P=0.006）， and boosted their killing function against tumors （P＜0.001）. Compared with the control group， in vivo study demonstrated that Cordyceps sinensis extract monotherapy effectively inhibited the growth of PD-L1-resistant tumors ［tumor growth inhibition value （TGI）=26.1%， P=0.090］， while its combination with anti-PD-L1 antibody significantly reversed PD-L1 resistance （TGI=50.6%， P＜0.001）. ConclusionThis study provides certain data support for the activation of anti-tumor immunity by Cordyceps sinensis extract， and offers a new treatment strategy for enhancing anti-tumor immunity and reversing immune resistance in lung cancer.

Background::Total human immunodeficiency virus (HIV) DNA and integrated HIV DNA are widely used markers of HIV persistence. Droplet digital polymerase chain reaction (ddPCR) can be used for absolute quantification without needing a standard curve. Here, we developed duplex ddPCR assays to detect and quantify total HIV DNA and integrated HIV DNA.Methods::The limit of detection, dynamic ranges, sensitivity, and reproducibility were evaluated by plasmid constructs containing both the HIV long terminal repeat (LTR) and human CD3 gene (for total HIV DNA) and ACH-2 cells (for integrated HIV DNA). Forty-two cases on stable suppressive antiretroviral therapy (ART) were assayed in total HIV DNA and integrated HIV DNA. Correlation coefficient analysis was performed on the data related to DNA copies and cluster of differentiation 4 positive (CD4 +) T-cell counts, CD8 + T-cell counts and CD4/CD8 T-cell ratio, respectively. The assay linear dynamic range and lower limit of detection (LLOD) were also assessed. Results::The assay could detect the presence of HIV-1 copies 100% at concentrations of 6.3 copies/reaction, and the estimated LLOD of the ddPCR assay was 4.4 HIV DNA copies/reaction (95% confidence intervals [CI]: 3.6-6.5 copies/reaction) with linearity over a 5-log 10-unit range in total HIV DNA assay. For the integrated HIV DNA assay, the LLOD was 8.0 copies/reaction (95% CI: 5.8-16.6 copies/reaction) with linearity over a 3-log 10-unit range. Total HIV DNA in CD4 + T cells was positively associated with integrated HIV DNA ( r = 0.76, P <0.0001). Meanwhile, both total HIV DNA and integrated HIV DNA in CD4 + T cells were inversely correlated with the ratio of CD4/CD8 but positively correlated with the CD8 + T-cell counts. Conclusions::This ddPCR assay can quantify total HIV DNA and integrated HIV DNA efficiently with robustness and sensitivity. It can be readily adapted for measuring HIV DNA with non-B clades, and it could be beneficial for testing in clinical trials.

Background::Cluster of differentiation 8 (CD8 T) cells play critical roles in eradicating human immunodeficiency virus (HIV)-1 infection, but little is known about the effects of T cells expressing CD8 at low levels (CD8 low) or high levels (CD8 high) on HIV-1 replication inhibition after HIV-1 invasion into individual. Methods::Nineteen patients who had been acutely infected with HIV-1 (AHI) and 20 patients with chronic infection (CHI) for ≥2 years were enrolled in this study to investigate the dynamics of the quantity, activation, and immune responses of CD3 +CD8 low T cells and their counterpart CD3 +CD8 high T cells at different stages of HIV-1 infection. Results::Compared with healthy donors, CD3 +CD8 low T cells expanded in HIV-1-infected individuals at different stages of infection. As HIV-1 infection progressed, CD3 +CD8 low T cells gradually decreased. Simultaneously, CD3 +CD8 high T cells was significantly reduced in the first month of AHI and then increased gradually as HIV-1 infection progressed. The classical activation of CD3 +CD8 low T cells was highest in the first month of AHI and then reduced as HIV-1 infection progressed and entered the chronic stage. Meanwhile, activated CD38 -HLA-DR +CD8 low T cells did not increase in the first month of AHI, and the number of these cells was inversely associated with viral load ( r = -0.664, P = 0.004) but positively associated with the CD4 T-cell count ( r = 0.586, P = 0.014). Increased programmed cell death protein 1 (PD-1) abundance on CD3 +CD8 low T cells was observed from the 1st month of AHI but did not continue to be enhanced, while a significant T cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibition motif (ITIM) domains (TIGIT) abundance increase was observed in the 12th month of infection. Furthermore, increased PD-1 and TIGIT abundance on CD3 +CD8 low T cells was associated with a low CD4 T-cell count (PD-1: r = -0.456, P = 0.043; TIGIT: r = -0.488, P = 0.029) in CHI. Nonetheless, the nonincrease in PD-1 expression on classically activated CD3 +CD8 low T cells was inversely associated with HIV-1 viremia in the first month of AHI ( r = -0.578, P = 0.015). Notably, in the first month of AHI, few CD3 +CD8 low T cells, but comparable amounts of CD3 +CD8 high T cells, responded to Gag peptides. Then, weaker HIV-1-specific T-cell responses were induced in CD3 +CD8 low T cells than CD3 +CD8 high T cells at the 3rd and 12th months of AHI and in CHI. Conclusions::Our findings suggest that CD3 +CD8 low T cells play an anti-HIV role in the first month of infection due to their abundance but induce a weak HIV-1-specific immune response. Subsequently, CD3 +CD8 low T-cell number decreased gradually as infection persisted, and their anti-HIV functions were inferior to those of CD3 +CD8 high T cells.

Objective:To observe the multimodal imaging characteristics of combined hamartoma of the retina and retinal pigment epithelium (CHRRPE).Methods:A cross-sectional study was conducted.Sixteen eyes of 16 patients with CHRRPE were enrolled in Second People's Hospital of Yunnan Province from March 2013 to July 2019.Fundus color photography, fundus autofluorescence (FAF), fluorescein fundus angiography (FFA), indocyanine green angiography (ICGA), optical coherence tomography (OCT), optical coherence tomography angiography (OCTA) and multicolor imaging were performed in all patients.The multimodal imaging characteristics were analyzed.This study protocol adhered to the Declaration of Helsinki and was approved by an Ethics Committee of Second People's Hospital of Yunnan Province (No.20130106). Written informed consent was obtained from patients or their guardians prior to any medical examination.Results:Tumors were located in the posterior optic disc, and translucent glial lesions with unclear borders and slight elevations were observed.The surface of the lesions was attached by different degrees of fibroproliferative membrane, and the adjacent vessels were twisted and dilated.The tumors presented flat bulging green reflexes on the retina at the posterior pole by multicolor imaging, and OCT image showed thickened optic disc and retina near the optic disc, structural disorder, high reflectance of the surface, and low reflectance of the deep retina below the periretinal membrane.OCTA showed irregular blood flow signals, and the signal of retinal blood vessels was twisted and dilated.FAF showed that the autofluorescence intensity of tumors was weakened to different degrees.Early lesions presented different degrees of blocked fluorescence in FFA.Deformed and tortuous blood vessels were found in the eyes, and telangiectasia showing needle-like punctate strong fluorescence leakage was observed in severe eyes by FFA.ICGA showed no abnormal choroidal vessels.Conclusions:The main imaging features of CHRRPE include abnormal retinal blood vessels in the tumor area and fibrous proliferative membranes on the surface in color image; OCT shows that the retina and the retinal pigment epithelium are involved, and the retina in the tumor is thickened with disordered structure; high reflection OCTA shows irregular internal blood flow signals inside the tumor; FFA examination shows fluorescence obscuration and obviously tortuous retinal blood vessels.Multimodal imaging examinations are helpful for the diagnosis of CHRRPE.

Objective:To assess the three-dimensional dose distribution in radiotherapy plans using the structural similarity index(SSIM), compare the performance of SSIM with commonly used quality assessment indices, and develop a SSIM-based quality assessment method of multiple prescribed doses.Methods:The SSIM was introduced to providea quality score of various voxels by comparing actual and ideal three-dimensional dose data and combining the spatial location information of the voxels. Then the average value in a region of interest (ROI) was calculated as the quality score of the region. Fifty-three cases of cervical cancer were selected to analyze the correlation of the SSIM with the uniformity index (HI), conformity index (CI) of the dose distribution in various ROIs and to explore the capability of the SSIM to reflect the uniformity and conformity of dose distribution.Two types of quality defects were individually introduced into two of 53 radiotherapy plans. Then the two plans were compared with normal plans to characterize the response of the SSIM.Results:There was no correlation between HI and SSIM in positive lymph nodes(PGTVnd) due to the decrease in the HI sensitivity, while there was a significant negative correlation between them in regions where PGTVnd was removed from the planning target volume(PTV, R=-0.86, P<0.01). Meanwhile, there was a significant positive correlation between CI and SSIM in PGTVnd ( R=0.83, P<0.01). Therefore, the SSIM can be used to identify the artificial design defects in plans by determining abnormal dose gradients. Conclusions:Apart from reducing the defects of previous assessment parameters, the SSIM has the capability to assess the quality of radiotherapy plans by combining the uniformity and conformity of dose distribution and can provide accurate feedback on the spatial locations of quality defects.

Objective:To study the influence of G protein-coupled estrogen receptor 1 (GPER1) specific agonist G1 and antagonist G2 in epilepsy susceptibility of temporal lobe epileptic rats.Methods:Sixty rats were randomly divided into control group, G1 treatment group and G15 treatment group ( n=20). Rats in the latter two groups were intraperitoneally injected with GPER1 agonist G1 (10 μg) or antagonist G15 (40 μg) for a consecutive 12 d. Lithium chloride pilocarpine epilepsy models were prepared in the 3 groups. The behavior manifestations of these rats were observed within 1 h of intraperitoneal injection of pilocarpine; Racine grading was used to evaluate the severity of epileptic seizures every 5 min; the latency of epileptic seizures (Racine grading IV) and epileptic seizure grading at different time points in the 3 groups were compared. The EEG monitoring was performed to these rats, and EEG data were recorded from 10 min before pilocarpine injection to 2 h after pilocarpine injection; EEG time-frequency was analyzed by Fast-Fourier transform (FFT); distribution of brain electrical energy and changes of θ and α wave energy during 20 min of epileptic status were compared among the 3 groups. Results:(1) As compared with that in the control group and G1 treatment group, the latency of epileptic seizures in the G15 treatment group was significantly shortened ( P<0.05); 15 and 20 min after pilocarpine injection, the epileptic seizure grading of rats in G1 treatment group was statistically lower than that in control group ( P<0.05); 15-35 min after pilocarpine injection, the epileptic seizure grading of rats in G15 treatment group was significantly higher than that in control group ( P<0.05). (2) As compared with those in the control group, rats in the G1 treatment group had smaller brain wave amplitude, while the rats in the G15 treatment group had earlier seizure time, larger brain wave amplitude and higher frequency. There were no obvious changes in the amount of brain electrical energy between the G1 treatment group and control group; while the amount of brain electrical energy in the G15 treatment group 2 h after pilocarpine injection was higher than that in the control group. As compared with those in the control group and G1 treatment group, the θ and α wave energy values of rats in the G15 treatment group were significantly increased within 20 min of epileptic status ( P<0.05). Conclusion:Activation level of GPER1 might be associated with susceptibility to epileptic seizures, and specific inhibition of GPER1 activation can enhance the susceptibility to epilepsy and increase the energy values of specific frequency bands during epilepsy.

Objective:To evaluate the effect of imatinib on growth impairment in children with chronic myeloid leukemia (CML-CP) in the chronic phase.Methods:From July 2018 to July 2019, questionnaires were distributed to CML children aged <18 years at the time of diagnosis who were receiving imatinib for at least 3 months or to their parents in China. The height-for-age standard deviation score (HtSDS) and the difference of standard deviation integral (△HtSDS) were used to explore the change in height with imatinib therapy.Results:The data of 238 respondents were included; 138 (58.0% ) respondents were men. The median age at the first diagnosis of CML was 11.0 years (range, 1.4-17.9 years) , and 93 (39.0% ) respondents were at the prepuberty stage. At the time of completing the questionnaires, the median age was 15.0 years (range, 2.0-34.0 years) . The median duration of imatinib therapy was 28 months (range, 3-213 months) . Among all the respondents, the mean HtSDS when completing the questionnaires (-0.063±1.361) was significantly lower than that at the time of starting imatinib treatment (0.391±1.244) ( P<0.001) . Total 71.0% respondents showed growth impairment that was more common in those starting imatinib therapy at prepubertal age than in those starting at pubertal age. Multivariate analysis showed that younger at the start of imatinib therapy ( P<0.001) and longer duration of imatinib therapy ( P<0.001) were significantly associated with severe growth impairment on imatinib therapy. Conclusions:Imatinib induced growth impairment in children with CML-CP. Younger the age of initiation and longer the duration of imatinib therapy, more obvious the effect of imatinib on growth impairment.

Objective: To compare the biomechanical performance between the single- versus double-threaded cannulated screws in the treatment of femoral neck fractures of Pauwels type Ⅲ. Methods: Models of femoral neck fracture of Pauwels type Ⅲ (70°) were made of the Sawbone synthetic composite femurs. All specimens were divided into 2 groups (n=12). Group A was fixated with single-threaded cannulated screws and group B with double-threaded cannulated screws, both in an inverted triangle configuration. The screws ranged from 90 to 95 mm in length and from 7.3 to 7.5 mm in diameter. All the specimens were subjected to axial stiffness and failure load tests with 7° valgus (simulating normal two-legged weight-bearing stance) and 25° valgus (simulating normal one-legged weight-bearing stance) and torsion test as well. The 2 groups were compared in the torques at axial stiffness angles of 1°, 2°, 3°, 5° and 7°. Results: Group B had significantly greater axial stiffness at 7° valgus and 25° valgus (89±26 N/mm and 128±37 N/mm) and failure load (1,154±368 N) than group A did (36±12 N/mm and 47±16 N/mm; 688±94 N) (P< 0.05). The torques increased with the increase in rotation angle in both groups. However, the torques in group B (3.26±0.96, 4.16±1.23, 4.64±1.13, 5.59±1.26 and 6.53±1.47 N·m) were all significantly larger than in group A (1.44±0.19, 2.03±0.41, 2.33±0.62, 2.74±0.87 and 3.05±1.07 N·m) (P<0.05). Conclusion Double-threaded cannulated screws may provide better biomechanical stability than single-threaded ones, due to their substantial improvement in anti-compression and anti-rotation performance.

Objective To study the effect of transcutaneous electrical acupoint stimulation (TEAS) on emergence agitation in children undergoing cochlear implantation.Methods Sixty patients (age range,1-4 years) scheduled to undergo pediatric cochlear implantation were enrolled and randomized into the TEAS group (group T,n =30) that received continuous TEAS and the control group (group C,n =30) that did not receive TEAS.Anesthesia time,operation time,change in blood flow dynamics during the perioperative period,extubation time,and PACU stay time were recorded.The incidence of emergence agitation and postoperative nausea and vomiting (PONV) was monitored.Results No significant intergroup differences were observed in age,sex,weight,operation time,and PACU stay time (P ＞ 0.05).The MAP and HR in group T were lower than those in group C at different time points such as 30 min after the start of surgery,the end of the surgery,the extubation and 5 min after extubation,and the difference was statistically significant (P ＜ 0.05).The FPS-R,FLACC,and PAED scores in group T were lower than those in group C at different time points within 30 min after exmbation (including the time of extubation),and the difference was statistically significant (P ＜ 0.05).Extubation time,anesthesia time,and incidence of PONV were lower in group T than in group C,and the difference was statistically significant (P ＜ 0.05).Conclusion TEAS combined with general anesthesia significantly reduces the incidence of emergence agitation in children undergoing surgery for congenital hearing loss;it also helps control hypotension,reheve dramatic hemodynamic changes during the waking period,and reduce the incidence of PONV.

Objective: To evaluate the efficacy and safety of maintenance therapy with reduced dose of rhTPO in the patients with primary immune thrombocytopenia (ITP) who attained stable platelet (PLT) counts after daily administration of rhTPO. Methods: Treatment was started with a daily administration of rhTPO (300 U/kg) for 2 consecutive weeks. Patients who attained stable PLT≥50×10⁹/L were enrolled to maintenance therapy starting with every other day administration of rhTPO, then adjusted dose interval to maintain platelet count (30-100) ×10⁹/L. Results: A total of 91 eligible patients were enrolled. Fourteen patients discontinued the study due to noncompliance (12/14) and investigator decision (2/14) . Among 77 patients who completed the study, 38 patients with the administration of rhTPO at every other day or less could maintain PLT≥30×10⁹/L for 12 weeks. The percentage of patients with a platelet response (PLT≥30×10⁹/L) at 4^th week, 8^th week and 12^th week of maintain therapy was 92.6% (63/68) , 82.7% (43/52) and 85.0% (34/40) , respectively. Median platelet counts remained in the range of (70-124) ×10⁹/L. The overall incidence of rhTPO-related adverse events was 7.7%. All the adverse events were generally mild. Conclusion Extending the dose interval of rhTPO is feasible to maintain stable platelet count in the patients with ITP, but the optimal dose interval is uncertain and might vary with individuals.