ObjectiveTo investigate the processing technology of calcined Haematitum based on the concept of quality by design（QbD） and to assess its health risk. MethodsTaking whole iron content， Fe²⁺ dissolution content and looseness as critical quality attributes（CQAs）， and calcination temperature， calcination time， spreading thickness and particle size as critical process parameters（CPPs） determined by the failure mode and effect analysis（FMEA）， the processing technology of calcined Haematitum was optimized by orthogonal test combined with analytic hierarchy process-criteria importance through intercriteria correlation（AHP-CRITIC） hybrid weighting method. The contents of heavy metals and harmful elements were determined by inductively coupled plasma mass spectrometry， and the health risk assessment was carried out by daily exposure（EXP）， target hazard quotient（THQ） and lifetime cancer risk（LCR）， and the theoretical value of the maximum limit was deduced. ResultsThe optimal processing technology for calcined Haematitum was calcination at 650 ℃， calcination time of 1 h， particle size of 0.2-0.5 cm， spreading thickness of 1 cm， and vinegar quenching for 1 time［Haematitum-vinegar（10：3）］. The contents of 5 heavy metals and harmful elements in 13 batches of calcined Haematitum were all decreased with reductions of up to 5-fold. The cumulative THQ of 2 batches of samples was>1， while the cumulative THQ of all batches of Haematitum was>1. The LCR of As in 1 batches of Haematitum was 1×10^-6-1×10^-4， and the LCR of the rest was<1×10^-6， and the LCRs of calcined Haematitum were all<1×10^-6， indicating that the carcinogenic risk of calcined Haematitum was low， but special attention should still be paid to Haematitum medicinal materials. Preliminary theoretical values of the maximum limits of Cu， As， Cd， Pb and Hg were formulated as 1 014， 25， 17， 27， 7 mg·kg^-1. ConclusionThe optimized processing technology of calcined Haematitum is stable and feasible， and the contents of heavy metals and harmful elements are reduced after processing. Preliminary theoretical values of the maximum limits of Cu， As， Cd， Pb and Hg are formulated to provide a scientific basis for the formulation of standards for the limits of harmful elements in Haematitum.

ObjectiveBased on the concept of quality by design（QbD）， the processing process of calcined Pyritum was optimized， and its X-ray diffraction（XRD） fingerprint was established. MethodsThe safety， effectiveness and quality controllability of calcined Pyritum were taken as the quality profile（QTPP）， the color， hardness， metallic luster， phase composition， the contents of heavy metals and hazardous elements were taken as the critical quality attributes（CQAs）， and the calcination temperature， calcination time， paving thickness and particle size were determined as the critical process parameters（CPPs）. Differential thermal analysis， X-ray diffraction（XRD） and inductively coupled plasma mass spectrometry（ICP-MS） were used to analyze the correlation between the calcination temperature and CQAs of calcined Pyritum. Then， based on the criteria importance through intercriteria correlation（CRITIC）-entropy weight method， the optimal processing process of calcined Pyritum was optimized by orthogonal test. Powder XRD was used to analyze the phase of calcined Pyritum samples processed according to the best process， and the mean and median maps of calcined Pyritum were established by the superposition of geometric topological figures， and similarity evaluation and cluster analysis were carried out. ResultsThe results of single factor experiments showed that the physical phase of Pyritum changed from FeS₂ to Fe₇S₈ during the process of temperature increase， the color gradually deepened from dark yellow， and the contents of heavy metals and harmful elements decreased. The optimized processing process of calcined Pyritum was as follows：calcination temperature at 750 ℃， calcination time of 2.5 h， paving thickness of 3 cm， particle size of 0.8-1.2 cm， vinegar quenching 1 time［Pyritum-vinegar（10∶3）］. After calcination， the internal structure of Pyritum was honeycomb-shaped， which was conducive to the dissolution of active ingredients. XRD fingerprints of 13 batches of calcined Pyritum characterized by 10 common peaks were established. The similarities of the relative peak intensities of the XRD fingerprints of the analyzed samples were>0.96， and it could effectively distinguish the raw products and unqualified products. ConclusionTemperature is the main factor affecting the quality of calcined Pyritum. After processing， the dissolution of the effective components in Pyritum increases， and the contents of heavy metals and harmful substances decrease， reflecting the function of processing to increase efficiency and reduce toxicity. The optimized processing process is stable and feasible， and the established XRD fingerprint can be used as one of the quality control standards of calcined Pyritum.

ObjectiveTo investigate the processing technology of calcined Haematitum based on the concept of quality by design（QbD） and to assess its health risk. MethodsTaking whole iron content， Fe²⁺ dissolution content and looseness as critical quality attributes（CQAs）， and calcination temperature， calcination time， spreading thickness and particle size as critical process parameters（CPPs） determined by the failure mode and effect analysis（FMEA）， the processing technology of calcined Haematitum was optimized by orthogonal test combined with analytic hierarchy process-criteria importance through intercriteria correlation（AHP-CRITIC） hybrid weighting method. The contents of heavy metals and harmful elements were determined by inductively coupled plasma mass spectrometry， and the health risk assessment was carried out by daily exposure（EXP）， target hazard quotient（THQ） and lifetime cancer risk（LCR）， and the theoretical value of the maximum limit was deduced. ResultsThe optimal processing technology for calcined Haematitum was calcination at 650 ℃， calcination time of 1 h， particle size of 0.2-0.5 cm， spreading thickness of 1 cm， and vinegar quenching for 1 time［Haematitum-vinegar（10：3）］. The contents of 5 heavy metals and harmful elements in 13 batches of calcined Haematitum were all decreased with reductions of up to 5-fold. The cumulative THQ of 2 batches of samples was>1， while the cumulative THQ of all batches of Haematitum was>1. The LCR of As in 1 batches of Haematitum was 1×10^-6-1×10^-4， and the LCR of the rest was<1×10^-6， and the LCRs of calcined Haematitum were all<1×10^-6， indicating that the carcinogenic risk of calcined Haematitum was low， but special attention should still be paid to Haematitum medicinal materials. Preliminary theoretical values of the maximum limits of Cu， As， Cd， Pb and Hg were formulated as 1 014， 25， 17， 27， 7 mg·kg^-1. ConclusionThe optimized processing technology of calcined Haematitum is stable and feasible， and the contents of heavy metals and harmful elements are reduced after processing. Preliminary theoretical values of the maximum limits of Cu， As， Cd， Pb and Hg are formulated to provide a scientific basis for the formulation of standards for the limits of harmful elements in Haematitum.

ObjectiveBased on the concept of quality by design（QbD）， the processing process of calcined Pyritum was optimized， and its X-ray diffraction（XRD） fingerprint was established. MethodsThe safety， effectiveness and quality controllability of calcined Pyritum were taken as the quality profile（QTPP）， the color， hardness， metallic luster， phase composition， the contents of heavy metals and hazardous elements were taken as the critical quality attributes（CQAs）， and the calcination temperature， calcination time， paving thickness and particle size were determined as the critical process parameters（CPPs）. Differential thermal analysis， X-ray diffraction（XRD） and inductively coupled plasma mass spectrometry（ICP-MS） were used to analyze the correlation between the calcination temperature and CQAs of calcined Pyritum. Then， based on the criteria importance through intercriteria correlation（CRITIC）-entropy weight method， the optimal processing process of calcined Pyritum was optimized by orthogonal test. Powder XRD was used to analyze the phase of calcined Pyritum samples processed according to the best process， and the mean and median maps of calcined Pyritum were established by the superposition of geometric topological figures， and similarity evaluation and cluster analysis were carried out. ResultsThe results of single factor experiments showed that the physical phase of Pyritum changed from FeS₂ to Fe₇S₈ during the process of temperature increase， the color gradually deepened from dark yellow， and the contents of heavy metals and harmful elements decreased. The optimized processing process of calcined Pyritum was as follows：calcination temperature at 750 ℃， calcination time of 2.5 h， paving thickness of 3 cm， particle size of 0.8-1.2 cm， vinegar quenching 1 time［Pyritum-vinegar（10∶3）］. After calcination， the internal structure of Pyritum was honeycomb-shaped， which was conducive to the dissolution of active ingredients. XRD fingerprints of 13 batches of calcined Pyritum characterized by 10 common peaks were established. The similarities of the relative peak intensities of the XRD fingerprints of the analyzed samples were>0.96， and it could effectively distinguish the raw products and unqualified products. ConclusionTemperature is the main factor affecting the quality of calcined Pyritum. After processing， the dissolution of the effective components in Pyritum increases， and the contents of heavy metals and harmful substances decrease， reflecting the function of processing to increase efficiency and reduce toxicity. The optimized processing process is stable and feasible， and the established XRD fingerprint can be used as one of the quality control standards of calcined Pyritum.

Immunotherapy has emerged as a revolutionary approach to treat immune-related diseases. Dendritic cells (DCs) play a pivotal role in orchestrating immune responses, making them an attractive target for immunotherapeutic interventions. Modulation of gene expression in DCs using genome editing techniques, such as the CRISPR-Cas system, is important for regulating DC functions. However, the precise delivery of CRISPR-based therapies to DCs has posed a significant challenge. While lipid nanoparticles (LNPs) have been extensively studied for gene editing in tumor cells, their potential application in DCs has remained relatively unexplored. This study investigates the important role of cholesterol in regulating the efficiency of BAMEA-O16B lipid-assisted nanoparticles (BLANs) as carriers of CRISPR/Cas9 for gene editing in DCs. Remarkably, BLANs with low cholesterol density exhibit exceptional mRNA uptake, improved endosomal escape, and efficient single-guide RNA release capabilities. Administration of BLAN_mCas9/gPD-L1 results in substantial PD-L1 gene knockout in conventional dendritic cells (cDCs), accompanied by heightened cDC1 activation, T cell stimulation, and significant suppression of tumor growth. The study underscores the pivotal role of cholesterol density within LNPs, revealing potent influence on gene editing efficacy within DCs. This strategy holds immense promise for the field of cancer immunotherapy, offering a novel avenue for treating immune-related diseases.

Objective To investigate the correlation between the expression of the activator of HSP90 ATPase-1(AHA1),lysyl oxidase like-2 protein(LOXL2)in osteosarcoma tissues with mRNA expression of invasion and metastasis genes and their clinical significance.Methods A total of 90 osteosarcoma patients diagnosed and treated in North China Medical and Health Group Fengfeng General Hospital from February 2016 to March 2017 were selected as the research object.The expression of AHA1 mRNA,LOXL2 mRNA and invasion and metastasis genes Wnt family member 9A(Wnt9a)mRNA,zinc finger E-box binding homologous box 1(ZEB1)mRNA,zinc finger E-box binding homologous box 1(ZEB2)mRNA,N-cadherin(N-cad)mRNA,and vimentin(Vim)mRNA in tissues were detected by real-time fluorescence quantitative PCR.Pearson correlation analysis was used for correlation analysis.The differences in expression of AHA1 mRNA and LOXL2 mRNA in osteosarcoma patients among different clinical characteristics were compared.Kaplan-Meier survival analysis was used to analyze the effect of AHA1 mRNA and LOXL2 mRNA on the prognosis of osteosarcoma patients.The prognostic factors of osteosarcoma patients were analyzed by univariate and multivariate COX regression.Results The expressions of AHA1 mRNA(3.16±0.59),LOXL2 mRNA(2.84±0.44)and invasion and metastasis genes[Wnt9a mRNA(3.23±0.42),ZEB1 mRNA(2.73±0.39),ZEB2 mRNA(2.52±0.56),N-cad mRNA(2.71±0.65)and Vim mRNA(2.81±0.73)]in osteosarcoma tissues were higher than those in paracancerous tissues(1.10±0.21,0.95±0.18,0.79±0.15,0.64±0.11,0.98±0.19,0.68±0.14,0.72±0.15),and the differences were statissically significant(t=31.206,37.716,51.903,48.931,24.706,28.964,26.605,all P<0.05).There was a significant positive correlation between AHA1 mRNA and LOXL2 mRNA expression in osteosarcoma(r=0.712,P<0.05).The expressions of AHA1 mRNA and LOXL2 mRNA were significantly positively correlated with the expressions of invasion and metastasis genes(Wnt9a,ZEB1,ZEB2,N-cad,and Vim mRNA)in tumor tissue of osteosarcoma group(r=0.504～0.720,all P<0.05).The expressions of AHA1 mRNA and LOXL2 mRNA in osteosarcoma tissues with Eneeking stage Ⅲ,soft tissue infiltration,and lung metastasis were higher than those in patients with Eneeking stage Ⅰ～Ⅱ,no soft tissue infiltration,and no lung metastasis,with significant differences(t=14.122～171.054,all P<0.05).The 5-year survival rates of patients in the AHA1 mRNA high expression group and low expression group were 36.36%(16/44)and 78.26%(36/46),respectively.The 5-year cumulative survival rate of patients in the AHA1 mRNA high expression group was significantly lower than that in the low expression group(Log-rank χ2=16.081,P<0.05).The 5-year survival rates of patients with high and low expression of LOXL2 mRNA were 34.88%(15/43)and 78.72%(37/47),respectively.The 5-year cumulative survival rate of patients in the LOXL2 mRNA high expression group was significantly lower than that in the low expression group(Log-rank χ2=15.880,P<0.05).Lung metastasis(OR=1.921,P<0.05),Eneeking stage Ⅲ(OR=1.906,P<0.05),AHA1 mRNA high expression(OR=1.405,P<0.05),and LOXL2 mRNA high expression(OR=1.733,P<0.05)were independent risk factors affecting the poor survival prognosis of osteosarcoma patients.Conclusion The expressions of AHA1 mRNA and LOXL2 mRNA in osteosarcoma were increased,and they were correlated with the expression of invasion and metastasis genes,indicating they may be independent risk factors affecting the poor survival and prognosis of osteosarcoma patients.

Grooming,as an evolutionarily conserved repetitive behavior,is common in various animals,including humans,and serves essential functions including,but not limited to,hygiene maintenance,thermoregulation,de-arousal,stress reduction,and social behaviors.In rodents,grooming involves a patterned and sequenced structure,known as the syntactic chain with four phases that comprise repeated stereotyped movements happening in a cephalocaudal progression style,beginning from the nose to the face,to the head,and finally ending with body licking.The context-dependent occurrence of grooming behavior indicates its adaptive significance.This review briefly summarizes the neural substrates responsible for rodent grooming behavior and explores its relevance in rodent models of neuropsychiatric disorders and neurodegenerative diseases with aberrant grooming phenotypes.We further emphasize the utility of rodent grooming as a reliable measure of repetitive behavior in neuropsychiatric models,holding promise for translational psychiatry.Herein,we mainly focus on rodent self-grooming.Allogrooming(grooming being applied on one animal by its conspecifics via licking or carefully nibbling)and heterogrooming(a form of grooming behavior directing towards another animal,which occurs in other contexts,such as maternal,sexual,aggressive,or social behaviors)are not covered due to space constraints.

Purpose To explore the value of dual-layer spectral detector CT(DLSCT)in evaluating preoperative tumor staging in rectal adenocarcinoma(RA).Materials and Methods A total of 78 patients with pathologically confirmed RA in Guangdong Provincial Hospital of Chinese Medicine from May 2021 to March 2022 were involved in this retrospective study.All the patients underwent plain and dual-phase contrast-enhanced scans by DLSCT within one week before surgery.Taking pathological results as the golden standard,the accuracy rates of tumor staging were calculated and compared between the multiple-parameter maps derived from DLSCT and 120 kVp polyenergetic image.The effective atomic number(Z-eff)from plain scan,iodine concentration(IC)from arterial phase(AP)and venous phase(VP)were measured.The normalized iodine concentration(NIC)in AP and VP were calculated.The differences of Z-eff,NICAP and NICVP were compared among the pT1-2,pT3 and pT4 groups.The correlation between the pT stages and above values was assessed and the diagnostic efficiencies were analyzed.Results The overall accuracy rate(88.46%vs.67.95%;χ2=9.628,P=0.002),the pT1-2 staging accuracy rate(80.00%vs.40.00%;χ2=6.667,P=0.01),and the pT3 staging accuracy rate(88.10%vs.69.05%;χ2=4.525,P=0.033)of multiple-parameter maps derived from DLSCT were significantly higher than those of 120 kVp polyenergetic image,respectively.The Z-eff,NICAP and NICVP were significantly different among pT stage groups(F=6.456,11.029,12.698,all P<0.05)and exhibited a positive correlation with pT stages(r=0.371,0.367,0.363,all P<0.01).The areas under the curve of Z-eff,NICAP and NICVP to assess pT3-4 staging RA were 0.77,0.71 and 0.74,respectively.Conclusion The multiple-parameter maps derived from DLSCT can significantly improve the diagnostic accuracy of preoperative tumor staging of RA.Z-eff from plain scan and NIC from dual-phase helps differentiate pT1-2 from pT3-4 staging RA.

Vaccination is a main measure for protecting chickens against Marek's disease,while it is not able to suppress the infection,proliferation,transmission,and virulence enhancement on Marek's disease virus.Inhibiting the proliferation of Marek's disease virus in chicken is therefore an im-portant option for enhancing defense effectiveness.In this study,a compound,DUBs-IN-1,was found to inhibit the activity of MDV049,a protease encoded by Marek's disease virus,via screening a protease inhibitor library using MDV049 as target and ubiquitin probe.Molecular docking re-vealed that DUBs-IN-1 can interact with the residues which formed the catalytic pocket of MDV049,blocking the interaction between Ub substrate and the catalytic center of MDV049,then suppress the activity of MDV049 with competitive inhibition.Using the CPE model,it was found that DUBs-IN-1 at the concentration of 0.35 and 0.70 μmol/L significantly inhibited the CPE in-duced by Marek's disease virus in CEF cells.Quantitative analysis revealed that DUBs-IN-1 inhibi-ted the proliferation of Marek's disease virus in CEF cells(P＜0.01).Furthermore,it was found that the administration of 80 and 150 pg/(kg·d)of DUBs-IN-1 in chicken infected by Marek's disease virus significantly inhibited the proliferation of MDV in T cells(P＜0.01).In summary,this study demonstrated that the compound DUBs-IN-1 is able to inhibit the proliferation of Marek's disease virus in chicken cells,laying a theoretical and practical foundation for further de-velopment of the drugs against Marek's disease virus.

Objective To study the eye irritation and the pharmacokinetics of tacrolimus-loaded cationic nanoemulsion-based in-situ gel in rabbits. Methods The eye irritation of tacrolimus-loaded cationic nanoemulsion-based in-situ gel in rabbits was observed by histological cross-sections of external ocular tissues stained with HE. The aqueous humor of rabbit eyes was extracted by corneal puncture and analyzed by HPLC-MS for pharmacokinetic study. Results Tacrolimus-loaded cationic nanoemulsion-based in-situ gel had no significant irritation on rabbit eyes. The pharmacokinetic parameter showed that the AUC of tacrolimus-loaded cationic nanoemulsion-based in-situ gel was (128.34±13.09) ng·h/ml, which was 1.13 times of tacrolimus-loaded cationic nanoemulsion (113.61±12.36) ng·h/ml and 1.88 times of Talymus® (68.25±10.82) ng·h /ml. Conclusion Tacrolimus-loaded cationic nanoemulsion-based in-situ gel had the advantages of low irritation, long retention time and high bioavailability in rabbit eyes. It has a good potential for clinical application.