BACKGROUND:Bone tissue loss caused by tumors and trauma can have an adverse effect on postoperative rehabilitation.Therefore,scaffold materials are usually implanted during treatment.However,the existing implant materials are relatively simple and lack antibacterial properties.Early implantation may lead to iatrogenic autoinfection and have an adverse effect on osteogenesis.OBJECTIVE:To construct a KR-12-a5 polypeptide-nanohydroxyapatite-small intestinal submucosa composite scaffold and evaluate its feasibility as a material for promoting bone defect repair.METHODS:The small intestinal submucosa scaffold and the small intestinal submucosa scaffold containing 25,50,and 100 mg/mL nanohydroxyapatite(referred to as nHA-SIS scaffold)were prepared by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride/N-hydroxysuccinimide cross-linking method.The appropriate scaffold was screened for subsequent experiments by mechanical property testing.The antibacterial properties of KR-12-a5 polypeptide solution against Staphylococcus aureus,Streptococcus gordonii,and Fusobacterium nucleatum were detected.The nHA-SIS scaffolds were immersed in 250,500,and 1 000 μg/mL KR-12-a5 peptide solutions for 24 hours,and then freeze-dried to obtain peptide-loaded nanohydroxyapatite-porcine small intestinal submucosa composite scaffolds(denoted as P-nHA-SIS scaffolds).The sustained-release properties of the three groups of scaffolds were characterized.The nHA-SIS scaffolds and the three groups of P-nHA-SIS scaffolds were co-cultured with Staphylococcus aureus,Streptococcus gordonii,and Fusobacterium nucleatum for 24 hours or 48 hours.The scaffolds with strong antibacterial ability were screened by live and dead bacteria staining and scanning electron microscopy for subsequent experiments.The degradation properties and water absorption rates of the uncross-linked small intestinal submucosa scaffolds,cross-linked small intestinal submucosa scaffolds,nHA-SIS scaffolds,and P-nHA-SIS scaffolds were characterized.The extracts of cross-linked small intestinal submucosal scaffolds,nHA-SIS scaffolds,and P-nHA-SIS scaffolds were co-cultured with MC3T3-E1 cells.CCK-8 assay and live-dead cell staining were performed.The effects of the extracts of the three scaffolds on the migration of MC3T3-E1 cells were detected by Transwell chamber assay.RESULTS AND CONCLUSION:(1)The elastic modulus and compressive strength of 25,50,and 100 mg/mL nHA-SIS scaffolds were higher than those of small intestinal submucosal scaffolds(P＜0.05),among which the elastic modulus and compressive strength of 25 mg/mL nHA-SIS scaffolds were the highest,and this group of scaffolds were selected for subsequent experiments to load peptides.(2)KR-12-a5 peptide had strong antibacterial activity against common bacteria in bone defects(Staphylococcus aureus,Streptococcus gordonii,and Fusobacterium nucleatum).The three groups of P-nHA-SIS scaffolds all had sustained release properties.With the increase of peptide mass concentration,the antibacterial property of P-nHA-SIS scaffold was enhanced.Among them,the P-nHA-SIS scaffold loaded with 500 μg/mL peptide had achieved a satisfactory antibacterial effect,and this group of scaffolds would be selected in the future.(3)The degradation rate of the three groups of cross-linked scaffolds was lower than that of the uncross-linked scaffolds,and the water absorption rate was greater than that of the uncross-linked scaffolds.P-nHA-SIS scaffolds could promote the proliferation and migration of MC3T3-E1 cells without affecting the activity of MC3T3-E1 cells.(4)The results show that P-nHA-SIS scaffolds have strong antibacterial properties and the ability to promote the proliferation and migration of MC3T3-E1 cells,and are expected to be used in bone defect repair.

BACKGROUND:Bone tissue loss caused by tumors and trauma can have an adverse effect on postoperative rehabilitation.Therefore,scaffold materials are usually implanted during treatment.However,the existing implant materials are relatively simple and lack antibacterial properties.Early implantation may lead to iatrogenic autoinfection and have an adverse effect on osteogenesis.OBJECTIVE:To construct a KR-12-a5 polypeptide-nanohydroxyapatite-small intestinal submucosa composite scaffold and evaluate its feasibility as a material for promoting bone defect repair.METHODS:The small intestinal submucosa scaffold and the small intestinal submucosa scaffold containing 25,50,and 100 mg/mL nanohydroxyapatite(referred to as nHA-SIS scaffold)were prepared by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride/N-hydroxysuccinimide cross-linking method.The appropriate scaffold was screened for subsequent experiments by mechanical property testing.The antibacterial properties of KR-12-a5 polypeptide solution against Staphylococcus aureus,Streptococcus gordonii,and Fusobacterium nucleatum were detected.The nHA-SIS scaffolds were immersed in 250,500,and 1 000 μg/mL KR-12-a5 peptide solutions for 24 hours,and then freeze-dried to obtain peptide-loaded nanohydroxyapatite-porcine small intestinal submucosa composite scaffolds(denoted as P-nHA-SIS scaffolds).The sustained-release properties of the three groups of scaffolds were characterized.The nHA-SIS scaffolds and the three groups of P-nHA-SIS scaffolds were co-cultured with Staphylococcus aureus,Streptococcus gordonii,and Fusobacterium nucleatum for 24 hours or 48 hours.The scaffolds with strong antibacterial ability were screened by live and dead bacteria staining and scanning electron microscopy for subsequent experiments.The degradation properties and water absorption rates of the uncross-linked small intestinal submucosa scaffolds,cross-linked small intestinal submucosa scaffolds,nHA-SIS scaffolds,and P-nHA-SIS scaffolds were characterized.The extracts of cross-linked small intestinal submucosal scaffolds,nHA-SIS scaffolds,and P-nHA-SIS scaffolds were co-cultured with MC3T3-E1 cells.CCK-8 assay and live-dead cell staining were performed.The effects of the extracts of the three scaffolds on the migration of MC3T3-E1 cells were detected by Transwell chamber assay.RESULTS AND CONCLUSION:(1)The elastic modulus and compressive strength of 25,50,and 100 mg/mL nHA-SIS scaffolds were higher than those of small intestinal submucosal scaffolds(P＜0.05),among which the elastic modulus and compressive strength of 25 mg/mL nHA-SIS scaffolds were the highest,and this group of scaffolds were selected for subsequent experiments to load peptides.(2)KR-12-a5 peptide had strong antibacterial activity against common bacteria in bone defects(Staphylococcus aureus,Streptococcus gordonii,and Fusobacterium nucleatum).The three groups of P-nHA-SIS scaffolds all had sustained release properties.With the increase of peptide mass concentration,the antibacterial property of P-nHA-SIS scaffold was enhanced.Among them,the P-nHA-SIS scaffold loaded with 500 μg/mL peptide had achieved a satisfactory antibacterial effect,and this group of scaffolds would be selected in the future.(3)The degradation rate of the three groups of cross-linked scaffolds was lower than that of the uncross-linked scaffolds,and the water absorption rate was greater than that of the uncross-linked scaffolds.P-nHA-SIS scaffolds could promote the proliferation and migration of MC3T3-E1 cells without affecting the activity of MC3T3-E1 cells.(4)The results show that P-nHA-SIS scaffolds have strong antibacterial properties and the ability to promote the proliferation and migration of MC3T3-E1 cells,and are expected to be used in bone defect repair.

AIM: To develop a novel gene-delivery therapeutic based on CRISPR/Cas9 genome editing technology capable of specifically targeting and knocking out the VEGFA gene, thereby achieving sustained suppression of VEGFA expression in retinal pigment epithelial(RPE)cells and providing a new strategy for gene therapy in retinal neovascular diseases.METHODS:Single guide RNAs targeting the human VEGFA gene for knockout were designed, and corresponding recombinant plasmids were constructed. A novel polymer(PTEE)was used to encapsulate the plasmids to prepare a PTEE-loaded anti-VEGFA plasmid(PLAP)gene delivery system. PTEE materials at concentrations of 0.1, 0.2, 0.4, 0.8, and 1.6 μg/μL were co-incubated with ARPE-19 cells, and the biocompatibility of PTEE was evaluated using the cell counting kit-8(CCK-8)assay. Recombinant plasmids expressing green fluorescent protein(GFP)were constructed. Lipofectamine 3000 and jetOPTIMUS®DNA transfection reagents were used as control groups, and PTEE nanomaterials were used as the experimental group to encapsulate the plasmids. When the cell confluence reached 80%, the formulations were transfected into ARPE-19 and 293T cells. GFP expression was observed under light microscopy, and the transfection efficiencies of each group were compared. ARPE-19 cells were induced under hypoxia, and PLAP was transfected into the cells. The expression level of VEGFA was detected by enzyme-linked immunosorbent assay(ELISA)to evaluate the efficacy of this novel gene delivery system.RESULTS: After co-incubation of ARPE-19 cells with different concentrations of PTEE for 24 h and 48 h, no significant effect on cell viability was observed in any group. The transfection efficiency of PLAP in ARPE-19 cells was higher than that in the Lipo3000 and jetOPTIMUS groups, with statistically significant differences(P<0.01). Hypoxia for 6 h significantly induced the upregulation of VEGFA mRNA expression in ARPE-19 cells, and under hypoxic conditions, the PTEE group exhibited a significant inhibitory effect on VEGFA expression(P<0.01).CONCLUSION:PLAP exhibits favorable biocompatibility and prominent VEGFA inhibitory effects in vitro, making it a potential candidate drug for gene therapy of retinal neovascular diseases.

Sj?gren's syndrome(SS)is one of the common rheumatic diseases in clinical practice.Modern medicine commonly uses drugs such as artificial tears,saliva,glucocorticoids,immunosuppressants,and biologics to control the condition.Clinical practice has shown that in addition to modern medical basic treatment,the use of traditional Chinese medicine(TCM)can help improve the clinical efficacy of SS.According to the symptoms and signs of Sj?gren's syndrome in TCM,it is classified as"dryness and obstruction",and the core pathogenesis of the disease is spleen deficiency and deficiency of body fluids.Subsequently,toxic and pathogenic factors gather,leading to the decline of internal organs.The initial causes are spleen damage,unstable barrier,and invasion of pathogenic factors.The core link is spleen dysfunction,insufficient body fluid,and dryness arising from it.Spleen deficiency generates evil,obstruction of qi,and lack of body fluids are the root causes of illness.The main treatment method is the"spleen strengthening method",which treats spleen deficiency,dampness and stagnation,and the body fluid is not distributed.The treatment focuses on strengthening the spleen and qi,supplementing the lungs and generating fluids.Spleen deficiency leads to loss of vitality,blood stasis obstructs blood vessels,and the treatment is to strengthen the spleen,soothe the liver,remove blood stasis,and unblock the orifices.The spleen yang is not vigorous,and qi transformation is impaired.The treatment is to invigorate the spleen and warm the stomach,promote yang circulation,and promote diuresis.

Sj?gren's syndrome(SS)is one of the common rheumatic diseases in clinical practice.Modern medicine commonly uses drugs such as artificial tears,saliva,glucocorticoids,immunosuppressants,and biologics to control the condition.Clinical practice has shown that in addition to modern medical basic treatment,the use of traditional Chinese medicine(TCM)can help improve the clinical efficacy of SS.According to the symptoms and signs of Sj?gren's syndrome in TCM,it is classified as"dryness and obstruction",and the core pathogenesis of the disease is spleen deficiency and deficiency of body fluids.Subsequently,toxic and pathogenic factors gather,leading to the decline of internal organs.The initial causes are spleen damage,unstable barrier,and invasion of pathogenic factors.The core link is spleen dysfunction,insufficient body fluid,and dryness arising from it.Spleen deficiency generates evil,obstruction of qi,and lack of body fluids are the root causes of illness.The main treatment method is the"spleen strengthening method",which treats spleen deficiency,dampness and stagnation,and the body fluid is not distributed.The treatment focuses on strengthening the spleen and qi,supplementing the lungs and generating fluids.Spleen deficiency leads to loss of vitality,blood stasis obstructs blood vessels,and the treatment is to strengthen the spleen,soothe the liver,remove blood stasis,and unblock the orifices.The spleen yang is not vigorous,and qi transformation is impaired.The treatment is to invigorate the spleen and warm the stomach,promote yang circulation,and promote diuresis.

Objective:To evaluate the repeatability of anterior segment biometry measurements obtained using ultrasound biomicroscopy (UBM) and the ArcScan Insight 100, and to compare the agreement between the two devices.Methods:A cross-sectional study was conducted.Seventy myopic patients (70 eyes) who underwent V4c implantable collamer lens implantation at Henan Eye Hospital from March to May 2023 were included.The ArcScan Insight 100 and UBM were used to measure the following parameters three times: angle-to-angle distance (ATA), sulcus-to-sulcus distance (STS), anterior chamber depth (ACD), crystalline lens rise (CLR), anterior chamber width (ACW), ciliary body inner diameter (CBID), anterior chamber angle (ACA), trabecular-ciliary angle (TCA) and maximum ciliary body thickness (CBTmax). The repeatability of anterior segment biological measurements obtained using the two devices was assessed by intraclass correlation coefficient (ICC). The consistency between the two instruments was evaluated by Bland-Altman consistency test.This study complied with the Declaration of Helsinki and was approved by the Ethics Committee of Henan Eye Hospital (No.HNEECKY-2021[13]). All patients understood the purpose and significance of this study and signed the informed consent form.Results:The repeatability of ATA, STS, ACD, CLR, ACW, CBID, ACA, TCA and CBTmax measured by UBM and ArcScan Insight 100 was good (all ICC>0.9). There was no significant difference in ACD, ACW, and ACA between the two instruments ( t=0.696, -1.025, -1.447; all P>0.05). ATA, STS, CLR and CBID measured by UBM were lower and TCA and CBTmax were higher than those measured by UBM ArcScan Insight 100, and the differences were statistically significant ( t=-8.586, -12.551, -4.481, -4.420, 4.535, 7.812; all P<0.05). The differences of ATA, STS, ACD, CLR, ACW, CBID and CBTmax between UBM and ArcScan Insight 100 were 0.38, 0.47, -0.01, 0.07, 0.3, 0.26 and -0.21 mm, respectively, with the 95% limits of agreement (LoA) of (-0.34, 1.10), (-0.15, 1.09), (-0.28, 0.26), (-0.20, 0.35), (-0.33, 0.93), (-0.71, 1.23) and (-0.64, 0.23)mm, respectively, which showed good coherence.The differences in ACA and TCA measurements were 2.26° and -7.81°, respectively, and the 95%LoA values were (-23.36°, 27.89°) and (-36.05°, 20.43°), respectively, with poor coherence.There was a strong positive correlation in ACD measurements measured by UBM and ArcScan Insight 100 ( r=0.827, P<0.05). There were moderate positive correlations in ATA, STS, CLR, ACW and CBID ( r=0.678, 0.749, 0.617, 0.765, 0.519; all P<0.05). There was no significant correlation in ACA, TCA and CBTmax ( r=0.270, 0.032, 0.178; all P>0.05). Conclusions:The repeatability of ArcScan Insight 100 and UBM in measuring anterior segment biological parameters is good.However, the consistency of ACA, TCA and CBTmax measured by the two instruments is poor and may be affected by self-regulation of the body.ATA, STS, ACD, CLR, ACW and CBID have good consistency and can be used interchangeably.

Nitric oxide synthase(NOS)and its product nitric oxide are involved in learning and memory functions.Increasing evidence shows that NOS plays an important role in the pathogenesis of Alzheimer's disease,influencing β-amyloid protein(Aβ)deposition,neuroinflammation,oxidative stress,abnormal microglia activation,synapse damage,autophagy,abnormal mitochondrial function of nerve cells,and cerebral hypoperfusion or vascular endothelial cell injury.This review summarizes the recent evidence for the role of NOS in the pathogenesis of Alzheimer's disease and provides new feasible targets for the prevention and treatment of Alzheimer's disease.

Objective:To evaluate the repeatability of anterior segment biometry measurements obtained using ultrasound biomicroscopy (UBM) and the ArcScan Insight 100, and to compare the agreement between the two devices.Methods:A cross-sectional study was conducted.Seventy myopic patients (70 eyes) who underwent V4c implantable collamer lens implantation at Henan Eye Hospital from March to May 2023 were included.The ArcScan Insight 100 and UBM were used to measure the following parameters three times: angle-to-angle distance (ATA), sulcus-to-sulcus distance (STS), anterior chamber depth (ACD), crystalline lens rise (CLR), anterior chamber width (ACW), ciliary body inner diameter (CBID), anterior chamber angle (ACA), trabecular-ciliary angle (TCA) and maximum ciliary body thickness (CBTmax). The repeatability of anterior segment biological measurements obtained using the two devices was assessed by intraclass correlation coefficient (ICC). The consistency between the two instruments was evaluated by Bland-Altman consistency test.This study complied with the Declaration of Helsinki and was approved by the Ethics Committee of Henan Eye Hospital (No.HNEECKY-2021[13]). All patients understood the purpose and significance of this study and signed the informed consent form.Results:The repeatability of ATA, STS, ACD, CLR, ACW, CBID, ACA, TCA and CBTmax measured by UBM and ArcScan Insight 100 was good (all ICC>0.9). There was no significant difference in ACD, ACW, and ACA between the two instruments ( t=0.696, -1.025, -1.447; all P>0.05). ATA, STS, CLR and CBID measured by UBM were lower and TCA and CBTmax were higher than those measured by UBM ArcScan Insight 100, and the differences were statistically significant ( t=-8.586, -12.551, -4.481, -4.420, 4.535, 7.812; all P<0.05). The differences of ATA, STS, ACD, CLR, ACW, CBID and CBTmax between UBM and ArcScan Insight 100 were 0.38, 0.47, -0.01, 0.07, 0.3, 0.26 and -0.21 mm, respectively, with the 95% limits of agreement (LoA) of (-0.34, 1.10), (-0.15, 1.09), (-0.28, 0.26), (-0.20, 0.35), (-0.33, 0.93), (-0.71, 1.23) and (-0.64, 0.23)mm, respectively, which showed good coherence.The differences in ACA and TCA measurements were 2.26° and -7.81°, respectively, and the 95%LoA values were (-23.36°, 27.89°) and (-36.05°, 20.43°), respectively, with poor coherence.There was a strong positive correlation in ACD measurements measured by UBM and ArcScan Insight 100 ( r=0.827, P<0.05). There were moderate positive correlations in ATA, STS, CLR, ACW and CBID ( r=0.678, 0.749, 0.617, 0.765, 0.519; all P<0.05). There was no significant correlation in ACA, TCA and CBTmax ( r=0.270, 0.032, 0.178; all P>0.05). Conclusions:The repeatability of ArcScan Insight 100 and UBM in measuring anterior segment biological parameters is good.However, the consistency of ACA, TCA and CBTmax measured by the two instruments is poor and may be affected by self-regulation of the body.ATA, STS, ACD, CLR, ACW and CBID have good consistency and can be used interchangeably.

People with AIDS are highly prone to opportunistic infections,among which pulmonary infections are the most common,with high incidence and fatality rates.How to effectively prevent or reduce the occurrence of HIV-related lung infections(HRLI)is of great practical significance.From the perspective of traditional Chinese medicine(TCM),HRLI and AIDS share common pathological mechanisms,including qi deficiency,spleen defi-ciency,phlegm-heat or phlegm-dampness obstructing the lungs.Based on the TCM theory of"preventing disease before it occurs",the three-level prevention approach includes:the first step"preventing disease before it occurs"emphasizes strengthening the body's defense mechanisms by tonifying the spleen and benefiting qi,and nourishing the lungs and kidneys to strengthen the exterior and protect the body;the second step"preventing the progression of disease once it occurs"emphasizes the removal of pathogenic factors while strengthening the body's defenses.Based on the principles of syndrome differentiation and treatment in TCM,focuses are kept on the spleen tonifying and phlegm resolving,pathogenic factors dispelling and dampness eliminating,while protecting the spleen and nourishing other organs.The third step"preventing relapse after recovery"emphasizes replenishing deficiencies and enhancing immunity.Adopting the three-level prevention approach and early intervention can reduce the inci-dence of HRLI,which is a new idea for the prevention and treatment of HRLI with TCM.

People with AIDS are highly prone to opportunistic infections,among which pulmonary infections are the most common,with high incidence and fatality rates.How to effectively prevent or reduce the occurrence of HIV-related lung infections(HRLI)is of great practical significance.From the perspective of traditional Chinese medicine(TCM),HRLI and AIDS share common pathological mechanisms,including qi deficiency,spleen defi-ciency,phlegm-heat or phlegm-dampness obstructing the lungs.Based on the TCM theory of"preventing disease before it occurs",the three-level prevention approach includes:the first step"preventing disease before it occurs"emphasizes strengthening the body's defense mechanisms by tonifying the spleen and benefiting qi,and nourishing the lungs and kidneys to strengthen the exterior and protect the body;the second step"preventing the progression of disease once it occurs"emphasizes the removal of pathogenic factors while strengthening the body's defenses.Based on the principles of syndrome differentiation and treatment in TCM,focuses are kept on the spleen tonifying and phlegm resolving,pathogenic factors dispelling and dampness eliminating,while protecting the spleen and nourishing other organs.The third step"preventing relapse after recovery"emphasizes replenishing deficiencies and enhancing immunity.Adopting the three-level prevention approach and early intervention can reduce the inci-dence of HRLI,which is a new idea for the prevention and treatment of HRLI with TCM.