Chinese hamster ovary (CHO) cells are the most established and versatile mammalian expression system for the large-scale production of recombinant therapeutic proteins, owing to their genetic stability, adaptability to serum-free suspension culture, and ability to perform human-like post-translational modifications. More than 70% of biologics approved by the U.S. Food and Drug Administration rely on CHO-based production platforms, underscoring their central role in modern biopharmaceutical manufacturing. Despite these advantages, CHO systems continue to face three persistent bottlenecks that limit their potential for high-yield, reproducible, and cost-efficient production: excessive metabolic burden during high-density culture, heterogeneity of glycosylation patterns, and progressive loss of long-term expression stability. This review provides an integrated analysis of recent advances addressing these challenges and proposes a forward-looking framework for constructing intelligent and sustainable CHO cell factories. In terms of metabolic regulation, excessive lactate and ammonia accumulation disrupts energy balance and reduces recombinant protein synthesis efficiency. Optimization of culture parameters such as temperature, pH, dissolved oxygen, osmolarity, and glucose feeding can effectively alleviate metabolic stress, while supplementation with modulators including sodium butyrate, baicalein, and S-adenosylmethionine promotes specific productivity (qP) by modulating apoptosis and chromatin structure. Furthermore, genetic engineering strategies—such as overexpression of MPC1/2, HSP27, and SIRT6 or knockout of Bax, Apaf1, and IGF-1R—have demonstrated significant improvements in cell viability and product yield. The combination of multi-omics metabolic modeling with artificial intelligence (AI)-based prediction offers new opportunities for building self-regulating CHO systems capable of dynamic adaptation to environmental stress. Regarding glycosylation uniformity, which determines therapeutic efficacy and immunogenicity, gene editing-based glycoengineering (e.g., FUT8 knockdown or ST6Gal1 overexpression) has enabled the humanization of CHO glycan profiles, minimizing non-human sugar residues and enhancing drug stability. Process-level strategies such as galactose or manganese co-feeding and fine control of temperature or osmolarity further allow rational regulation of glycosyltransferase activity. Additionally, in vitro chemoenzymatic remodeling provides a complementary route to construct human-type glycans with defined structures, though industrial applications remain constrained by cost and scalability. The integration of model-driven process design and AI feedback control is expected to enable real-time prediction and correction of glycosylation deviations, ensuring batch-to-batch consistency in continuous biomanufacturing. Long-term expression stability, another critical challenge, is often impaired by promoter silencing, chromatin condensation, and random genomic integration. Molecular optimization—such as the use of improved promoters (CMV, EF-1α, or CHO endogenous promoters), Kozak and signal peptide refinement, and incorporation of chromatin-opening elements (UCOE, MAR, STAR)—helps maintain durable transcriptional activity, while site-specific integration systems including Cre/loxP, Flp/FRT, φC31, and CRISPR/Cas9 can enable single-copy, position-independent gene insertion at genomic safe-harbor loci, ensuring stable, predictable expression. Collectively, this review highlights a paradigm shift in CHO system optimization driven by the convergence of genome editing, synthetic biology, and artificial intelligence. The transition from empirical optimization to rational, data-driven design will facilitate the development of programmable CHO platforms capable of autonomous regulation of metabolic flux, glycosylation fidelity, and transcriptional activity. Such intelligent cell factories are expected to accelerate the transformation from laboratory-scale research to industrial-scale, high-consistency, and economically sustainable biopharmaceutical manufacturing, thereby supporting the next generation of efficient and customizable biologics manufacturing.

Chinese hamster ovary (CHO) cells are the most established and versatile mammalian expression system for the large-scale production of recombinant therapeutic proteins, owing to their genetic stability, adaptability to serum-free suspension culture, and ability to perform human-like post-translational modifications. More than 70% of biologics approved by the U.S. Food and Drug Administration rely on CHO-based production platforms, underscoring their central role in modern biopharmaceutical manufacturing. Despite these advantages, CHO systems continue to face three persistent bottlenecks that limit their potential for high-yield, reproducible, and cost-efficient production: excessive metabolic burden during high-density culture, heterogeneity of glycosylation patterns, and progressive loss of long-term expression stability. This review provides an integrated analysis of recent advances addressing these challenges and proposes a forward-looking framework for constructing intelligent and sustainable CHO cell factories. In terms of metabolic regulation, excessive lactate and ammonia accumulation disrupts energy balance and reduces recombinant protein synthesis efficiency. Optimization of culture parameters such as temperature, pH, dissolved oxygen, osmolarity, and glucose feeding can effectively alleviate metabolic stress, while supplementation with modulators including sodium butyrate, baicalein, and S-adenosylmethionine promotes specific productivity (qP) by modulating apoptosis and chromatin structure. Furthermore, genetic engineering strategies—such as overexpression of MPC1/2, HSP27, and SIRT6 or knockout of Bax, Apaf1, and IGF-1R—have demonstrated significant improvements in cell viability and product yield. The combination of multi-omics metabolic modeling with artificial intelligence (AI)-based prediction offers new opportunities for building self-regulating CHO systems capable of dynamic adaptation to environmental stress. Regarding glycosylation uniformity, which determines therapeutic efficacy and immunogenicity, gene editing-based glycoengineering (e.g., FUT8 knockdown or ST6Gal1 overexpression) has enabled the humanization of CHO glycan profiles, minimizing non-human sugar residues and enhancing drug stability. Process-level strategies such as galactose or manganese co-feeding and fine control of temperature or osmolarity further allow rational regulation of glycosyltransferase activity. Additionally, in vitro chemoenzymatic remodeling provides a complementary route to construct human-type glycans with defined structures, though industrial applications remain constrained by cost and scalability. The integration of model-driven process design and AI feedback control is expected to enable real-time prediction and correction of glycosylation deviations, ensuring batch-to-batch consistency in continuous biomanufacturing. Long-term expression stability, another critical challenge, is often impaired by promoter silencing, chromatin condensation, and random genomic integration. Molecular optimization—such as the use of improved promoters (CMV, EF-1α, or CHO endogenous promoters), Kozak and signal peptide refinement, and incorporation of chromatin-opening elements (UCOE, MAR, STAR)—helps maintain durable transcriptional activity, while site-specific integration systems including Cre/loxP, Flp/FRT, φC31, and CRISPR/Cas9 can enable single-copy, position-independent gene insertion at genomic safe-harbor loci, ensuring stable, predictable expression. Collectively, this review highlights a paradigm shift in CHO system optimization driven by the convergence of genome editing, synthetic biology, and artificial intelligence. The transition from empirical optimization to rational, data-driven design will facilitate the development of programmable CHO platforms capable of autonomous regulation of metabolic flux, glycosylation fidelity, and transcriptional activity. Such intelligent cell factories are expected to accelerate the transformation from laboratory-scale research to industrial-scale, high-consistency, and economically sustainable biopharmaceutical manufacturing, thereby supporting the next generation of efficient and customizable biologics manufacturing.

Objective To analyze the effects of intravenous tirofiban following endovascular treatment on the prognosis of patients with acute anterior circulation large vessel occlusion stroke who did not achieve successful recanalization.Methods This retrospective study included consecutive patients with acute anterior circulation ischemic stroke who underwent endovascular treatment but did not achieve successful recanalization.These patients were retrospectively enrolled from the Department of Neurology at Nanjing Jinling Hospital,Affiliated Hospital of Medical School,Nanjing University,the Second Affiliated Hospital of Soochow University,Wuhan No.1 Hospital,and Yijishan Hospital of Wannan Medical College(the First Affiliated Hospital of Wannan Medical College)between January 2015 and April 2023.Baseline and clinical data were collected including age,sex,medical history(hypertension,diabetes,atrial fibrillation,hyperlipidemia),personal history(smoking and drinking),National Institutes of Health stroke scale(NIHSS)score at admission,trial of Org 10172 in acute stroke treatment(TOAST)classification(large artery atherosclerosis,cardioembolism,or other types),Alberta stroke program early CT score(ASPECTS)on admission,intravenous thrombolysis,onset-to-puncture time(OTP),collateral circulation status(poor collaterals:American Society of Interventional and Therapeutic Neuroradiology/Society of Interventional Radiology[ASTIN/SIR]score 0-2;good collateral:ASTIN/SIR score 3-4),procedure duration(from femoral or radial artery puncture to device removal),occlusion site,number of passes for thrombus retrieval,and post-procedural modified thrombolysis in cerebral infarction(mTICI)grade.Patients were divided into a tirofiban group and a non-tirofiban group based on whether they received post-procedural intravenous tirofiban.The primary outcome is a favorable functional outcome at 90-day since onset(modified Rankin scale[mRS]score ≤ 3).Safety outcomes included mortality at 90-day since onset(mRS score 6),any intracranial hemorrhage within 24 h post-procedure,and symptomatic intracranial hemorrhage within 24 h post-procedure.1∶1 propensity score matchings using the nearest neighbor method was performed based on variables with P＜0.05 in the comparison between the tirofiban and non-tirofiban groups.Differences in primary and safety outcomes between the matched groups were compared.Univariate and multivariate Logistic regression analyses were conducted with favorable outcomes at 90 days as the dependent variable to evaluate the effect of intravenous tirofiban administration after endovascular therapy on 90-day prognosis in patients without successful recanalization.Results A total of 356 patients without successful recanalization after endovascular treatment were included,comprising 195 males and 161 females,with a median age of 70(61,78)years(32-92 years).Among them,76 and 280patients were assigned to the tirofiban and non-tirofiban groups,respectively.At 90 days,114 patients had favorable outcomes,while 242 had unfavorable outcomes.(1)Before the 1∶1 propensity score matching,significant differences were observed between the tirofiban and non-tirofiban groups in terms of admission NIHSS score,ASPECTS,good collateral circulation,intravenous thrombolysis,and mTICI grade(all P＜0.05).After 1∶1 propensity score matching,66 matched pairs(132 patients)were obtained.No significant differences in the baseline and clinical characteristics were found between the two groups after matching(all P＞0.05).(2)After 1∶1 matching,a significant difference was observed in the rate of favorable outcomes at 90 days between the two groups(48.5％[32/66]vs.30.3％[20/66],P=0.033)after propensity score matching,while no significant differences were found in 90-day mortality,intracranial hemorrhage within 24h,or symptomatic intracranial hemorrhage within 24h(all P＞0.05).(3)Univariate analysis after 1∶1 matching indicated that age,atrial fibrillation,smoking history,admission NIHSS score,M2 segment middle cerebral artery occlusion,good collateral circulation,number of passes for thrombus retrieval,and tirofiban treatment were factors influencing favorable outcomes at 90 days.Multivariate Logistic regression analysis showed that younger age(aOR,0.942,95％CI 0.906-0.978,P=0.002),lower admission NIHSS score(aOR,0.855,95％CI 0.777-0.941,P=0.001),good collateral circulation(aOR,5.534,95％CI 2.141-14.301,P＜0.01),tirofiban treatment(aOR,2.774,95％CI 1.092-7.046,P=0.032),and M2 segment MCA occlusion(reference:internal carotid artery occlusion;aOR,4.874,95％CI 1.428-16.632,P=0.011)were independent predictors of favorable outcomes at 90 days.Conclusions Intravenous tirofiban administration after endovascular therapy may improve 90-day neurological outcomes in patients with acute anterior circulation large vessel occlusion stroke who did not achieve successful recanalization without increasing the risk of hemorrhage.Further large-scale randomized controlled trials are warranted to validate these findings.

The intelligent development of the medical field is burgeoning,and smart healthcare has become a crucial driver for enhancing medical service efficiency and improving patient experience.However,the specific pathways through which smart healthcare improves patient experience have not been fully explained.To address this gap,this study utilizes micro-level individual data collected by the Beijing Hospitals Authority in 2023 from patients in municipal hospitals,employing the Technology Acceptance Model(TAM)to investigate the impact mechanisms of smart service construction on patient experience.The results indicate that smart services significantly improve patient experience by enhancing perceived ease of use,with this effect being particularly prominent during the treatment stage.Additionally,smart services demonstrate potential advantages in alleviating the"digital divide,"with a more pronounced optimization effect on the healthcare experience of elderly populations.Highly educated groups exhibit greater satisfaction and perceived benefits from smart services.

An assay was established for detection of toxigenic Clostridioides difficile by combining microfluidic chip analysis with immunochromatography,and its performance was evaluated and compared with those of the Xpert C.difficile/Epi and VIDAS CD AB tests.Primer pairs were designed according to the tcdB and tpi genes in C.difficile.The specificity,limit of detection,reproducibility,and stability were evaluated.A total of 215 stool samples from patients with diarrhea were collected and tested in parallel with the Xpert C.difficile/Epi,VIDAS CDAB,and our assay.C.difficile was isolated from samples,and the tcdB gene was identified when discrepant results were obtained from the three above assays.Our assay showed no cross-reaction with other diarrhea-associated pathogens.Its reproducibility was 100％in testing of two standard plasmids containing tcdB and tpi genes at two concentrations(105 and 102 copies/μL).Two standard plasmids were detected after the PCR and immunochromatography reagents had been stored for 3,6,9,and 12 months,and all the results were posi-tive.The limit of detection was 10 copies/μL for toxigenic C.difficile.Testing of 33 samples positive for C.difficile with our assay(33/215,15.3％)yielded findings statistically coherent with those of the Xpert C.difficile/Epi test(kappa value=0.965).The sensitivity,specificity,positive predictive value,and negative predictive value of our assay,with respect to Xpert C.difficile/Epi as the standard,were 94.3％,100.0％,100.0％,and 98.9％;these values were significantly higher than those of VIDAS CDAB(60.0％,98.9％,91.3％,and 92.7％)(Kappa=0.714,OR=157.50,95％CI:62.03-847.28,P=0.013).In conclusion,our newly developed assay is specific,stable,and reproducible,and may be used for rapid and accu-rate detection of toxigenic C.difficile.The assay could be used for C.difficile infection screening in outpatient and emergen-cy,community medical service center,and epidemiological settings.

The intelligent development of the medical field is burgeoning,and smart healthcare has become a crucial driver for enhancing medical service efficiency and improving patient experience.However,the specific pathways through which smart healthcare improves patient experience have not been fully explained.To address this gap,this study utilizes micro-level individual data collected by the Beijing Hospitals Authority in 2023 from patients in municipal hospitals,employing the Technology Acceptance Model(TAM)to investigate the impact mechanisms of smart service construction on patient experience.The results indicate that smart services significantly improve patient experience by enhancing perceived ease of use,with this effect being particularly prominent during the treatment stage.Additionally,smart services demonstrate potential advantages in alleviating the"digital divide,"with a more pronounced optimization effect on the healthcare experience of elderly populations.Highly educated groups exhibit greater satisfaction and perceived benefits from smart services.

An assay was established for detection of toxigenic Clostridioides difficile by combining microfluidic chip analysis with immunochromatography,and its performance was evaluated and compared with those of the Xpert C.difficile/Epi and VIDAS CD AB tests.Primer pairs were designed according to the tcdB and tpi genes in C.difficile.The specificity,limit of detection,reproducibility,and stability were evaluated.A total of 215 stool samples from patients with diarrhea were collected and tested in parallel with the Xpert C.difficile/Epi,VIDAS CDAB,and our assay.C.difficile was isolated from samples,and the tcdB gene was identified when discrepant results were obtained from the three above assays.Our assay showed no cross-reaction with other diarrhea-associated pathogens.Its reproducibility was 100％in testing of two standard plasmids containing tcdB and tpi genes at two concentrations(105 and 102 copies/μL).Two standard plasmids were detected after the PCR and immunochromatography reagents had been stored for 3,6,9,and 12 months,and all the results were posi-tive.The limit of detection was 10 copies/μL for toxigenic C.difficile.Testing of 33 samples positive for C.difficile with our assay(33/215,15.3％)yielded findings statistically coherent with those of the Xpert C.difficile/Epi test(kappa value=0.965).The sensitivity,specificity,positive predictive value,and negative predictive value of our assay,with respect to Xpert C.difficile/Epi as the standard,were 94.3％,100.0％,100.0％,and 98.9％;these values were significantly higher than those of VIDAS CDAB(60.0％,98.9％,91.3％,and 92.7％)(Kappa=0.714,OR=157.50,95％CI:62.03-847.28,P=0.013).In conclusion,our newly developed assay is specific,stable,and reproducible,and may be used for rapid and accu-rate detection of toxigenic C.difficile.The assay could be used for C.difficile infection screening in outpatient and emergen-cy,community medical service center,and epidemiological settings.

Based on the results of the National Drug Sampling and Inspection Programme,we summarized the history,the standard collection,the production enterprises and the dosage form specifications,the quality standard study,the pharmacological and pharmacodynamic study,and the clinical application study of Wuwei Qingzhuo preparation of Mongolian medicine and Shiliu Jianwei preparation of Zang medicine,to provide the basis for improved quality standards for both preparations.The development of these two preparations was searched and analyzed through literature.The available information shows that there is very little research on the two preparations and insufficient pharmacological experimental and clinical experimental data.The two preparations are basically the same in prescription and efficacy.However,the quality standards are very different,which are not conducive to the quality control of the two and their related dosage forms.And it is suggested that the Chinese Pharmacopoeia should take the situation of this category into comprehensive consideration,and unify the quality standards of the two preparations.

Objective To investigate the clinical efficacy and safety of facilitated percutaneous coronary intervention(PCI)with half-dose recombinant staphylokinase(r-SAK)in patients with ST-segment elevation myocardial infarction(STEMI)who are expected to undergo PCI within 120 minutes.Methods From October 2021 to August 2022,a total of 200 STEMI patients in eight centers were included and randomly assigned in a 1﹕1 ratio to either r-SAK group or control group.Patients received loading doses of aspirin and ticagrelor and intravenous heparin and were randomized to receive an intravenous bolus of either 5 mg r-SAK or normal saline prior to PCI.The outcomes were set as ST-segment resolution(STR)at 60-90 minutes after PCI,the proportion and transition of pathological Q waves on the 5th day after PCI,and the proportion of high-sensitivity cardiac troponin T(hs-cTnT)peaking within 12 hours of onset.The safety outcome was major bleeding events defined as Bleeding Academic Research Consortium(BARC)≥type 3 bleeding during hospitalization.Results Compared with the control group,the r-SAK group had a higher proportion of STR≥70%within 60-90 minutes after PCI(58.3%vs.40.3%,P=0.009);a lower proportion of pathological Q waves(59.1%vs.74.1%,P=0.040);a lower rate of Q wave progression(14.8%vs.43.2%,P＜0.001);a higher rate of Q wave disappearance(12.5%vs.3.7%,P=0.027);and a higher proportion of hs-cTnT peaking within 12 hours of symptom onset[31/40(77.5%)vs.17/33(51.5%),P=0.027].Regarding the safety outcome,no significant difference in BARC≥type 3 bleeding was found between the two groups during hospitalization(P＞0.05).Conclusions For STEMI patients who were expected to undergo primary PCI within 120 minutes of symptom onset,the facilitated PCI with half-dose r-SAK significantly increased the proportion of STR≥70%at 60-90 minutes after PCI,reduced the formation of pathological Q waves,and shortened the time to peak hs-cTnT,without increasing the risk of bleeding,which should be an alternative reperfusion strategy worthy of further study.

Objective To investigate the clinical efficacy and safety of facilitated percutaneous coronary intervention(PCI)with half-dose recombinant staphylokinase(r-SAK)in patients with ST-segment elevation myocardial infarction(STEMI)who are expected to undergo PCI within 120 minutes.Methods From October 2021 to August 2022,a total of 200 STEMI patients in eight centers were included and randomly assigned in a 1﹕1 ratio to either r-SAK group or control group.Patients received loading doses of aspirin and ticagrelor and intravenous heparin and were randomized to receive an intravenous bolus of either 5 mg r-SAK or normal saline prior to PCI.The outcomes were set as ST-segment resolution(STR)at 60-90 minutes after PCI,the proportion and transition of pathological Q waves on the 5th day after PCI,and the proportion of high-sensitivity cardiac troponin T(hs-cTnT)peaking within 12 hours of onset.The safety outcome was major bleeding events defined as Bleeding Academic Research Consortium(BARC)≥type 3 bleeding during hospitalization.Results Compared with the control group,the r-SAK group had a higher proportion of STR≥70%within 60-90 minutes after PCI(58.3%vs.40.3%,P=0.009);a lower proportion of pathological Q waves(59.1%vs.74.1%,P=0.040);a lower rate of Q wave progression(14.8%vs.43.2%,P＜0.001);a higher rate of Q wave disappearance(12.5%vs.3.7%,P=0.027);and a higher proportion of hs-cTnT peaking within 12 hours of symptom onset[31/40(77.5%)vs.17/33(51.5%),P=0.027].Regarding the safety outcome,no significant difference in BARC≥type 3 bleeding was found between the two groups during hospitalization(P＞0.05).Conclusions For STEMI patients who were expected to undergo primary PCI within 120 minutes of symptom onset,the facilitated PCI with half-dose r-SAK significantly increased the proportion of STR≥70%at 60-90 minutes after PCI,reduced the formation of pathological Q waves,and shortened the time to peak hs-cTnT,without increasing the risk of bleeding,which should be an alternative reperfusion strategy worthy of further study.