Previous studies found the dysbiosis of intestinal microbiota in diabetic kidney disease (DKD), especially the decreased SCFA‐producing bacteria. We aimed to investigate the concentration of the stool and serum short‐chain fatty acids (SCFAs), gut microbiota‐derived metabolites, in individuals with DKD and reveal the correlations between SCFAs and renal function.A total of 30 participants with DKD, 30 participants with type 2 diabetes mellitus (DM), and 30 normal controls (NC) in HwaMei Hospital were recruited from 1/1/2018 to 12/31/2019. Participants with DKD were divided into low estimated glomerular filtration rate (eGFR) and high eGFR  subgroups. Stool and serum were measured for SCFAs with gas chromatograph‐mass spectrometry.p<0.001, p<0.001, p=0.018, respectively) [1027.32(784.21–1357.90)]vs[2064.59(1561.82–2637.44)]μg/g,[929.53(493.65–1344.26)]vs[1684.57(1110.54–2324.69)]μg/g,[851.39(409.57–1611.65)] vs[1440.74(1004.15–2594.73)]μg/g, respectively, and the lowest fecal total SCFAs concentration among the groups. DKD group also had a lower serum caproate concentration than that with diabetes (p=0.020)[0.57(0.47–0.61)]vs[0.65(0.53–0.79)]μmol/L. In the univariate regression analysis, fecal and serum acetate correlated with eGFR . The correlation between serum total SCFAs and eGFR showed statistical significance  unadjusted and a borderline significance  when adjusted for Hb and LDL. The decrease in serum acetate and total SCFAs were found of borderline significant difference in both subgroups .The DKD group showed markedly lower levels of fecal acetate, propionate, and butyrate versus NC  were lowered, and there was a negative correlation between SCFAs and renal function. In this study,we measured the concentrations of fecal and serum SCFAs in the participants with DKD and it showed that serum and fecal SCFAs levels  were lowered, and there was a negative correlation between SCFAs and renal function. A recent study indicated that the prevalence of DM in China was 11.2% , especially in Han ethnicity.g_Escherichia‐Shigella and g_ Prevotella_9).Recent studies highlighted the involvement of gut‐kidney axis in nephropathy.via gut‐kidney axis, which also participated in kidney injury process. Being one of the major metabolites of microbiota‑mediated fiber fermentation process in the gut, short‐chain fatty acids (SCFAs) have attracted considerable interest. SCFAs are a subset of fatty acids that contain 6 or less carbon molecules and have shown beneficial effects on kidney.Besides, studies also indicated that gut microbiota and kidney were interacted In this study, all 90 participants were included from HwaMei Hospital. Fecal and serum samples were measured for SCFAs with gas chromatograph‐mass spectrometry (GC‐MS). We reported the substantial variations in the levels of fecal and serum SCFAs among normal controls, participants with diabetes, and participants with DKD. SCFA levels in participants with diabetic kidney disease were further analyzed within subgroups by renal function.22.1Scr formula. Participants in the NC group from physical examination center were given tests including metabolic panel, urinalysis, stool test, HBsAg (hepatitis B surface antigen), and anti‐HCV (hepatitis C antibody). Exclusions include: receiving antibiotics, probiotics, taking laxatives, or yogurt within 2 months, gastrointestinal or systemic diseases known to affect gut bacterial composition, primary or other secondary kidney diseases, obesity, liver cirrhosis with/without complications, non‐alcoholic fatty liver disease, HBsAg, or anti‐HCV positive. The clinical parameters are shown in Table There were 30 participants with DKD, 30 participants with type 2 diabetes, and 30 normal controls included in HwaMei Hospital, University of Chinese Academy of Science from January 1, 2018 to December 31, 2019. The diagnosis of T2DM was defined by the criterion issued by American Diabetes Association (ADA) in 2017.2.2Fresh fecal samples were collected and a portion of 200mg was utilized for each test. Blood samples were collected in the fasting status and serum was obtained by centrifugation at 3,500rpm for 5min at 4℃. These samples were then stored at −80°C until usage. One fecal sample and one serum sample in DKD group were later found not usable and were excluded in the study. 30 serum samples in NC group were not collected from the physical examination center. Hence, 30 fecal samples in NC group, 30 fecal and serum samples in the diabetes group, and 29 fecal and serum samples in the group with DKD were used for data determination.2.3Each fecal sample of 200mg was mixed with 0.8mL of ultrapure water, crushed with a tissue grinder, and then centrifuged at 12,000 rpm for 20 min at 4℃. Each 0.4ml supernatant was mixed with 0.1mL of 50% sulfuric acid (ultrapure water diluted), 0.5ml of ether  for 1 min, centrifuged at 12,000 rpm for 20 min at 4℃, and then stood for 30 min at 4℃. The supernatant ether layer was filtered through anhydrous sodium sulfate for GC–MS analysis.Each serum sample (100μL) was mixed with 50μL of 50% sulfuric acid (ultrapure water diluted), 200μL of ether  for 1 min, centrifuged at 12,000 rpm for 20 min at 4℃, and then stood for 30 min at 4℃. The supernatant ether layer was filtered through anhydrous sodium sulfate and the solution later transferred to a glass vial for GC–MS analysis.2.4The analysis was performed using the GC‐MS 7890A‐5975C . A FFAP capillary column (30m×0.25mm×0.25μm) was used for chromatographic separation, and helium (1 mL/min) was used as the carrier gas. The stepwise chromatographic thermal conditions were as follows: 100°C for 1 min, 5°C/min to 160°C, 40°C/min to 240°C, maintaining for 10 min. The mass spectrometer was set to scan mode at m/z 100–300 and selected ion monitoring mode at m/z 60 for acetate, butyrate, iso‐valerate, valerate, and caproate, maintaining for 4.72min、7.34min、8.90min、8.03min, and 11.26min respectively, as well as m/z 73 for propionate and iso‐butyrate for 5.90min and 6.31min separately.2.5p<0.1 in the univariate regression analysis were chosen for multivariate regression analysis. A P value<0.05 was considered statistically significant.All statistical analyses were performed with SPSS Statistics 19.0  and GraphPad Prism 7.0. The results were expressed as means with standard deviation (SD) for normally distributed continuous variables, median values (interquartile ranges) for non‐normally distributed continuous variables, and frequencies and percentages for categorical variables. ANOVA or Student's t‐test for independent samples was used for normally distributed continuous variables. Comparisons of non‐normally distributed continuous variables were performed using the Mann‐Whitney U‐test or Kruskal‐Wallis test. For categorical variables, the chi‐square test was used. Correlation difference between variables was analyzed by Spearman's R coefficient using psych package 1.9.12, and visualized by heatmap in corrplot package 0.84. The association between fecal or serum level with the clinic index was examined via binary logistic regression analysis, based on median level of fecal or serum SCFAs. Covariates with 33.1Baseline clinical and biochemical characteristics of all participants in NC group, DM group, and DKD group are shown in Table 3.2p=0.003) and NC group (p<0.001). Lower propionate and butyrate levels in DKD group were observed compared with NC group (p<0.05). Correspondingly, fecal total SCFAs presented in the same trend, being lowest in DKD group, 3843.01 (2491.81–5290.88) μg/g, while highest in NC group, reaching 6482.68 (4438.91–8379.59) μg/g (p<0.001). However, the median levels of iso‐butyrate, valerate, iso‐valerate, and caproate were equivalent among the three groups (p>0.05).The acetate, propionate, butyrate, iso‐butyrate, valerate, iso‐valerate, and caproate in stool sample were identified Figure . Notablyp<0.05). In addition, the differences of the concentrations of serum iso‐butyrate, valerate, and iso‐valerate between DKD group and DM group were approaching statistical significance, which were lower in DKD group . Apart from this, other SCFAs between DM group and DKD group showed no difference. Unexpectedly, there was no correlation between serum SCFAs and corresponding fecal SCFAs (raw p>0.05).Meanwhile, serum SCFAs were also measured in DM and DKD groups Figure . We obse3.3p=0.03; r=−0.27, p<0.01; r=−0.21, p=0.03, respectively). Meanwhile, UACR was negatively related to fecal acetate . Interestingly, hemoglobin and serum albumin levels showed a positive relationship with fecal acetate, propionate, and butyrate (p<0.05). Blood glucose was negatively related to fecal acetate and propionate .Correlations between the fecal SCFAs and clinical indicators were estimated by Spearman's correlation analysis Figure . As expep=0.04), positive correlations between total cholesterol, low‐density lipoprotein, and propionate .We further investigated the correlations between serum SCFAs and biochemical indicators Figure . Unexpecp=0.072; OR=1.017, 95%CI , p=0.032] (Tables p>0.05). Total SCFAs correlated with eGFR in subjects with statistical significance  unadjusted while the correlation became borderline significant  , p =0.049; OR=1.027, 95%CI , p=0.038] , 3.4p>0.05). UACR, serum creatinine, and blood urea nitrogen were higher (p<0.05) in the low GFR subgroup compared with the high GFR subgroup with statistically significant difference.To study the fecal and serum SCFAs in patients with various renal function, we categorized the DKD patients into two subgroups according to the eGFR level, the low GFR subgroup , and the high GFR subgroup . The baseline data of the two subgroups were shown in Table p>0.05). As shown in Table p=0.055, p=0.050, respectively). However, other SCFAs had no difference between these two subgroups (p>0.05).There were no differences in fecal SCFAs between the two subgroups , the major members of SCFAs, ameliorates mesangial matrix expansion, fibrosis, and inflammation in the kidneys of STZ‑induced diabetic rats.In this study, we did not use nutrition diaries, but all participants were interviewed for dietary habits and were explicitly asked for special dietary habits. Since all participants reported a Chinese omnivorous diet without any special dietary habits, dietary habits were unlikely to be a major confounder in the investigated subjects. However, there are some limitations in our cross‐section study, consequently we could not demonstrate the causal relationship between fecal, serum SCFAs, and the presence of DKD. This monocentric study included a small number of patients in China, prudence needs to be taken when trying to extrapolate our data to other populations. Besides, the composition and construction of gut microbiota in participants were not analyzed, therefore the relationship between fecal and serum SCFAs and gut microbiota was not identified.In conclusion, this study provides evidence for quantitative reduction of gut microbial products‐SCFAs  in DKD patients, demonstrating the association of SCFAs with renal function in DKD.The authors declare that they have no conflicts of interest.Study conception and design: KC, CZ, QL; Administrative support: KC, QL. Patient education and instruction: KC, CZ, LW, JL, WG, QZ, JZ; Collection and assembly of data: KC, CZ, ZD, LC, LW, JL, WG, QZ, JZ, CX, HL; Data analysis and interpretation: KC, CZ, QL; K and CZ took the lead in writing/wrote the manuscript with input from all authors. Manuscript writing: All authors. Final approval of manuscript: All authors.Figure S1Click here for additional data file.Figure S2Click here for additional data file.Table S1‐S2Click here for additional data file.


The Acrysof Cachet® angle-supported phakic intraocular lens (pIOL)  is designed to correct high refractive errors in human eyes. The aim of this study was to evaluate the outcome of AcrySof Cachet® angle-supported pIOL implantation with particular regard to efficacy and safety of the implant over a 60-month follow-up period.  Retrospective consecutive clinical case study.  Prior to pIOL implantation, patients had a complete ophthalmologic examination including objective and subjective refraction, uncorrected visual acuity (UCVA) and corrected distance visual acuity (CDVA), endothelial cells density (ECD), slit lamp photography, optical coherence tomography (OCT), Scheimpflug digital videokeratoscopy, optical biometry, slit lamp examination, intraocular pressure (IOP) measurement, and pupillometry. Postoperatively, patients received yearly a complete eye examination.  Thirty-one eyes of 16 patients were included in this study. The mean age was 36.2 ± 8.1 years. UCVA (logMAR) improved from 1.33 ± 0.20 before surgery to 0.08 ± 0.14 one year after surgery and was 0.20 ± 0.20 five years after surgery. CDVA (logMAR) improved from 0.10 ± 0.10 before surgery to 0.05 ± 0.13 one year after surgery and was 0.04 ± 0.14 five years postoperatively. The mean percentage of endothelial cells loss (ECL) was 11.51% over the first year and 15.95% five years after surgery. There were no intraoperative complications in any of the eyes.  Our results up to five years after implantation of the AcrySof Cachet® angle-supported pIOL demonstrated very good outcomes in all above shown measurements, including CDVA, UCVA, and ECD. However, since major endothelial cell loss may occur in some patients with this type of pIOL, regular follow-up visits are required. Phakic intraocular lenses (pIOLs) have been used for more than 60 years for correction of moderate to high ametropia , 2. HoweThe AcrySof Cachet® angle-supported pIOL (Alcon Laboratories) is a single-piece, foldable, soft acrylic lens (acrylate/methacrylate copolymer) with a 6.00 mm diameter optic and with an overall length of 12.5 to 14.0 mm and a dioptric range of −6.00 to –16.50 D in 0.5 D steps , 18. MadThirty-one eyes of 16 patients, who required surgical correction of high myopia to achieve spectacle independence between June 2010 and July 2012 at the Department of Ophthalmology, University Medical Center Mainz, were included in this study. The mean age of the patients was 36.19 ± 8.093 years, with a range from 23 to 50 years. The mean preoperative value of refraction, spherical equivalent, was −9.669 ± 2.730 D . The mean lens power of implanted IOLs was −10.240 ± 2.422 D, ranging from −6.000 to −15.500 D.2, mesopic pupil size <7.5 mm, anterior chamber depth (ACD) >3.0 mm, wish of spectacle independence, and ametropia not sufficiently correctable with excimer laser surgery.The inclusion criteria were age ≥18 years, stable refraction for at least 1 year, endothelial cells density (ECD) >2200 cells/mm2), anomaly of the iris or pupil, active infectious disease, recurrent or chronic uveitis, clinically significant cataract, posterior segment pathologies, such as macular degeneration or other macular and retinal abnormalities, and glaucoma.Contraindications for implantation included ACD <3.0 mm, insufficient ECD (<2200/mm  Uncorrected (UCVA) and corrected distance visual Scheimpflug acuity (CDVA)  Subjective and objective refraction  Measurement of ECD Prior to pIOL implantation, patients received a complete ophthalmologic evaluation including(1)Corneal topography and corneal thickness(2)ACD(3)  Axial length and ACD measurement   Slit lamp and a fundus examination   Measurement of IOP   Pupillometry Angle-to-angle distance.Scheimpflug digital videokeratoscopy  was performed with the measurement of  UCVA and CDVA  Subjective and objective refraction  Measurement of ECD  Anterior segment photography Postoperative follow-up visits were scheduled at day 1, week 1, and month 1, 3, 6, 12, 24, 36, 48, and 60 including the following examinations at given points of time:Position of the implant in the anterior chamber;Position of the haptics in the angle;Distance between the posterior surface of the pIOL and the anterior surface of the crystalline lens.Distance between the anterior surface of the pIOL and the central corneal endothelium;ACD;Scheimpflug digital videokeratoscopy was used to measureSlitlamp assessment and a fundus examinationMeasurement of IOP (Goldmann applanation tonometry)We used a questionnaire developed at our department to assess patient satisfaction after pIOL implantation.Anterior segment optical coherence tomography  was performed to measureSurgical procedure and postoperative treatment: surgery was performed under topical anesthesia, implanting one pIOL into one eye per session. All surgeries were performed by the same experienced ophthalmic surgeon (UV) following the manufacturer's recommendations. The pIOL was implanted through a 3.2 mm incision using the proprietary shooter system. Pupil constriction was obtained by topical administration of 0.5% pilocarpine eye drops and endothelial protection was ascertained using Viscoat® . The implantation process followed exactly the scheme as proposed by the manufacturer. Intraoperative viscoelastic removal was performed with bimanual irrigation/aspiration handpieces with low bottle height (infusion pressure approximately 25 mmHg). Complete removal was verified meticulously. Two hours postoperatively, all patients were subject to slit lamp examination and applanation tonometry of intraocular pressure. In all cases, readings were in physiological range without need of surgical revision or application of IOP-lowering medication. Two hours after surgery, the patient was visited to ensure complete removal of the viscoelastic material by IOP measurement.Postoperative treatment contained of unpreserved ofloxacin eye drops 4 times daily for 5 days and prednisolone eye drops 3 times daily for 3 weeks. The patients received yearly a complete eye examination including the above given measurements and also a photograph to control position, rotational stability, and pupil geometry .t-test and α = 0.05). For comparisons of postoperative values with the preoperative value, a paired t-test was used. Since each of the five postoperative time points was compared with the preoperative value, a correction for multiple comparisons was made by the Bonferroni correction, which resulted in a reduction of critical α to 0.01.Power calculation was conducted for ECL based on previously published data . For the∗∗∗p < 0.0001 for each year compared to preoperative values, p values were p=0.0339 (one year after surgery versus preoperative), p=0.0293 (two years after surgery versus preoperative), ∗p=0.0014 (three years after surgery versus preoperative), ∗p=0.0085 (four years after surgery versus preoperative), and p=0.0322 (five years after surgery versus preoperative) (UCVA (logMAR) improved from 1.33 ± 0.19 before surgery to 0.08 ± 0.14 one year after surgery and was 0.20 ± 0.20 five years after surgery (erative) . In none2 preoperatively, 2436 ± 503 cells/mm2 one year postoperatively, and 2314 ± 531 cells/mm2 five years postoperatively, which corresponds to an endothelial cell loss (ECL) of 11.51% over the first year and of 15.95% after 5 years , ∗p=0.0034 (two years after surgery versus preoperative), ∗p=0.0022 (three years after surgery versus preoperative), ∗p=0.0064 (four years after surgery versus preoperative), and ∗p=0.0071 (five years after surgery versus preoperative). The percentage of the eyes with an ECL of 25% or greater of preoperative ECD was 6.45% (2 of 31 eyes) [ECD was 2753 ± 322 cells/mm 5 years . Respect31 eyes) . In all p values were p=0.6454 (one year after surgery versus preoperative), p=0.2247 (two years after surgery versus preoperative), p=0.1509 (three years after surgery versus preoperative), p=0.3523 (four years after surgery versus preoperative), and p=0.0991 (five years after surgery versus preoperative).The mean IOP was 14.00 ± 2.39 mmHg before surgery, 14.36 ± 2.11 mmHg one year after surgery, and 18.92 ± 3.52 mmHg 5 years postoperatively. No marked IOP changes were observed during the whole postoperative period . RespectThere were no intraoperative complications in any of the eyes. However, in the first 24 hours postoperatively, two patients had a slightly elevated IOP of 23 mmHg, which dropped back to normal levels without any therapy within the following days. In one patient (43 years old when operated), there was a profound ECL of 63.82% after 5 years. The anterior chamber situation was normal without any inflammation. The distance between the pIOL and the lens as well as the position of the pIOL in the anterior chamber was stable, and the rotation was <5°.Because of the marked ECL, we decided to explant the Cachet® pIOL with immediate sequential implantation of an ICL®.Another patient (31 years old when operated), who had undergone Cachet® pIOL implantation in both eyes, experienced an IOP elevation up to 56 mmHg and 35 mmHg on the right and left eye, respectively, in the 5-years follow-up. After topical therapy with timolol/dorzolamide eye drops 2 times daily and clonidine eye drops 3 times daily for both eyes, the IOP values were between 13 mmHg and 20 mmHg in the right eye and between 12 mmHg and 19 mmHg in the left eye. The anterior chamber situation was normal without any inflammation in this patient. Rotation of the Cachet® pIOL in both eyes was <5°. The distance between the pIOL and the lens on the left eye was significantly smaller than on the right eye, but gonioscopy showed an open, normally appearing anterior chamber angle (Schaffer grade 3). Fundoscopic exam did not reveal any glaucomatous cupping of the optic nerve head. The patient is in regular outpatient examination, and the IOP is within normal limits with the therapy reported above. One patient (35 years old when operated), who received Cachet® pIOL bilaterally, was not satisfied with the postoperative UCVA. Therefore, we decided to perform femtosecond-assisted laser in-situ keratomileusis (Femto-LASIK) ten months after pIOL implantation in both eyes. The refraction after pIOL implantation, which needed correction was sph. −0.75 D cyl −0.25 D/ 77° on the right eye and sph. −0.5 D cyl. −0.5 D/ 35° on the left eye. In none of the patients signs of pigmentary dispersion  have been found in the follow-up examinations. Acute postoperative anterior uveitis was not observed. Pupil ovalization was not observed; pupil shape and motility were preserved in all patients. No iris synechiae, transillumination defects, or sphincter erosions were found. No opacities of the crystalline lens or decrease of the crystalline lens transparency could be found in any patient. There was no incidence of retinal detachment.n = 15 of 16, Three patients reported on night halos, two patients on increased light sensitivity at night, and one patient on glare 1 year postoperatively. With time, patients seemed to get used to these symptoms and it was not significant at the 5-year follow-up. As part of a satisfaction questionnaire, patients were asked if they would have the same pIOL implanted again, and at the 5-year postoperative visit, this question was positively answered by 94% (Our study demonstrated a high efficacy of Cachet® angle-supported pIOL implantation in correction of high refractive errors. We observed an increase in both UCVA and CDVA, which is in line with previous studies , 43. We Damage to the anterior chamber structures, especially to the corneal endothelium by the pIOL is the most specific complication. In a 7-year follow-up study, Alio et al. revealed an ECL of 3.8% in the first year, but of only 0.5% per year after the second postoperative year . Kohnen Pupil ovalization is regarded as a result of excessive continuous pressure upon the anterior chamber angle from the haptics. It is not only a cosmetically disturbing problem, but also causes glare. Alio et al. reported on pupil ovalization in 5.9% of patients, two of whom needed pIOL explantation . Perez-SIn conclusion, the AcrySof Cachet® pIOL showed excellent results in our retrospective study. There was only one case of IOL explantation because of marked ECL, which is of similar extent as in the recently published long-term analysis of Kohnen et al. . Our res


Three different LED spectra  with similar photosynthetic photon flux density (PPFD) were designed to explore the effects of supplementary far-red and blue lights on leaf color, biomass and phytochemicals of two cultivars of red-leaf lettuce (“Yanzhi” and “Red Butter”) in an artificial lighting plant factory. Lettuce plants under WB had redder leaf color and significantly higher contents of pigments, such as chlorophyll a, chlorophyll b, chlorophyll (a + b) and anthocyanins. The accumulation of health-promoting compounds, such as vitamin C, vitamin A, total phenolic compounds, total flavonoids and anthocyanins in the two lettuce cultivars were obviously enhanced by WB. Lettuce under WFR showed remarkable increase in fresh weight and dry weight; meanwhile, significant decreases of pigments, total phenolic compounds, total flavonoids and vitamin C were found. Thus, in the plant factory system, the application of WB can improve the coloration and quality of red leaf lettuce while WFR was encouraged for the purpose of elevating the yield of lettuce. Lactuca sativa L.) is one of the most common vegetables in PFALs because it is suitable for hydroponic growth and has a short growth period. Lettuce is popular as a salad vegetable, not only due to its tender taste, sweet and refreshing flavor and ornamental morphology with vivid colors, but also the abundant nutritional and antioxidant compounds such as vitamin C, sugars, proteins and phenolics [Indoor farms, especially plant factories with artificial light (PFALs), can produce high-value crops and increase crop production in a stable, controllable and safer manner . Lettucehenolics .Different light conditions  evoke different morphogenetic and photosynthetic responses in plants and the responses vary among plant species ,4,5. BluC. denticulatum plants [Far-red light (beyond 700 nm) has long been considered ineffective in photosynthesis for its poor absorption by mesophyll cells. It is known to induce shade-avoidance syndrome in plants, with the traits of increased plant height, reduced stem diameter and leaf area, narrower leaf angle and accelerated flowering ,12. Far-m plants . Previoum plants ,17,19,20As mentioned above, different lights and their combinations drive differential changes in plant architecture, photosynthesis and biosynthesis of phytochemicals. The light source is entirely artificial light in PFAL systems, which allows precise studies of effects of a particular light quality on plants by changing a narrow band of spectra. Therefore, in this study, the equalized-white LED lights whose band was basically consistent with PAR were used as control, the changes of leaf coloration, biomass and phytochemicals of two cultivars of red leaf lettuce in response to supplementary far-red light and blue light under similar PPFD were detected. Meanwhile, this study applied the heat map analysis and principal component analysis to achieve overall data visualization and simple clustering, which helped to fully evaluate the effects of supplementary light on lettuce. This study aimed to provide reference for the efficient LED light model with high productivity in PFAL systems.Lactuca sativa cv. “Yanzhi”, ShouheSeed, China) and red butterhead lettuce  were soaked for 1 h, then sowed in a moist sponge block (2 cm × 2 cm × 2 cm) and kept in the dark germination chamber. After 72 h, the germinated seeds with sponge blocks were grown in the DFT with half-strength nutrient solution. The full-strength nutrient solutions (EC ≈ 1.2 mS·cm−1 and pH ≈ 6.4) were composed of the following elements: 56.0 mg·L−1 N, 22.8 mg·L−1 P, 184.7 mg·L−1 K, 80.0 mg·L−1 Ca, 24.0 mg·L−1 Mg, 64 mg·L−1 S, 2.8 mg·L−1 Fe, 0.5 mg·L−1 B, 0.5 mg·L−1 Mn, 0.05 mg·L−1 Zn, 0.02 mg·L−1 Cu, 0.01 mg·L−1 Mo, temperature 21 ± 2 ℃, CO2 concentration 400–600 μmol·mol−1, relative humidity 55–60%, and equalized-white LED lighting at 250 μmol·m−2·s−1 PPFD from 8:00 to 18:00. After 18 days, the seedlings with three expanded true leaves were transplanted into the planting plate  with full-strength nutrient solution.This experiment was carried out in the PFAL, South China Agriculture University . The PFAL was equipped with a six-layer vertical hydroponic system using the deep flow technique . Each layer accommodated 15 planting plate and was divided into five cultivation regions whose light spectra can be adjusted independently. Seeds of red oakleaf lettuce (−2·s−1 (10 h·day−1 from 8:00 to 18:00). There were three light regimes  with equalized-white (peaking at 440 nm and 660 nm), far-red (730 ± 10 nm) and blue (450 ± 10 nm) LEDs were used as light sources. LED lightings at PPFD of 250 μmol·m regimes : W . The notation L* means lightness; a* represents the color from green to red; b* suggests the color from blue to yellow.v/v) solution and kept in darkness, 4 ℃ for 24 h until the leaf tissue turned white. Then the filtrates were measured at 645 nm, 663 nm and 440 nm by a UV-spectrophotometer . Contents of chlorophyll a (Chl a), chlorophyll b (Chl b), chlorophyll (a + b) (Chl (a + b)) and carotenoids (Car) were calculated according to the following equations [V is the volume of extract solution (mL), and W is the fresh weight (g) of the sample.Fresh tissue (0.5 g) from the mature leaves of lettuce was extracted with acetone-ethanol . These extractions were centrifuged at 12,000 rpm, 4 ℃ for 5 min. Then the supernatants were determined at 510 nm by using the UV-spectrophotometer. The TA values were calculated with the following equation:A1 is the absorbance of the sample extracted from pH 1.0 buffer solution, A2 is the absorbance of the sample extracted from pH 4.5 buffer solution. The number 484.8 represents the molecular weight of cyaniding-3-glucoside chloride. The dilution factor in this measurement is 1. The number 24.825 was the absorption coefficient at 510 nm.The total anthocyanin (TA) measurement was analyzed as previously described by Rapisarda et al. . Two groNine plants of lettuce in each treatment were randomly selected to measure the fresh and dry weight. The fresh weight (FW) was determined by analytical balance within 5 min upon harvest. Then these samples were oven-dried at 70 ℃ for 60 h to determine the corresponding dry weight (DW).The soluble proteins (SP) were determined according to Blakesley and Boezi . Fresh lv/v) and activated carbon powder (10 mg), and 80 ℃ water bath for 40 min each time. The mixture was filtered and then diluted to a total volume of 25 mL with 80% ethanol. Then 0.2 mL extract was mixed with 0.8 mL diluted water and 5 mL sulfuric acid anthrone reagent and 100 ℃ water bath for 10 min. The absorbance was detected at 625 nm by the UV-spectrophotometer.The soluble sugars (SS) were analyzed following the method described by Kohyama and Nishinari . Fresh lw/v, dissolved in H2SO4) reagent. Ten minutes later, 9.5 mL 8% NaOH (w/v) was added. The absorbance was measured at 410 nm with the UV-spectrophotometer.The nitrates were determined with the method proposed by Cataldo et al. . Fresh lFresh lettuce samples (0.5 g) were soaked in 8 mL methanol for 30 min in darkness. After 15 min centrifugation at 3000 rpm, the supernatant was kept in darkness, 4 ℃ for the measurements of DPPH radical inhibition percentage (DPPH) and ferric ion reducing antioxidant power (FRAP).iA: Supernatant of 2 mL mixed with 2 mL 0.2 μM DPPH; jA: Supernatant of 2 mL mixed with 2 mL ethanol; cA: 0.2 μM DPPH mixed with 2 mL ethanol) and determined at 517 nm by the UV-spectrophotometer. The DPPH radical inhibition percentage was calculated as following:The DPPH measurement was based on the method of Musa et al. . Three tv/v/v). The supernatant (0.4 mL) was added to the FRAP working reagent (3.7 mL), and the mixture was preserved in a 37 ℃ water bath for 10 min. The absorbance was determined at 593 nm by the UV-spectrophotometer. FeSO4·7H2O was used as the standard, and the results were expressed as mmol·g−1 FW.The FRAP measurement was according to Tadolini et al. . The FRAFresh lettuce samples (0.5 g) were soaked in 8 mL methanol for 30 min in darkness. After 15 min centrifugation at 3000 rpm, the supernatant was kept in darkness, 4 ℃ for the measurements of total phenolic compounds (TPC) and total flavonoids (TF).v/v) and 1.5 mL 26.7% Na2CO3 solution (w/v). The mixture was diluted to a total volume of 10 mL with ultra-pure water. After 2 h of reaction, the absorbance was recorded at 760 nm with the UV-spectrophotometer. TPC values were calculated from the gallic acid standard curve, and the results were expressed as mg gallic acid equivalent fresh weight (mgGAE·g−1 FW).The TPC measurement was conducted as stated by Tadolini et al. . The supw/v) and 0.7 mL NaNO2 solution (w/v). After 5 min, 0.7 mL 10% Al(NO3)3 solution (w/v) was added in for 5 min’ reaction. Then, 5 mL 5% NaOH solution (w%) and 8.6 mL 30% ethanol were added. The absorbance was determined at 510 nm with the UV-spectrophotometer. Rutin hydrate was used as the standard, and the results were expressed as mg·g−1 FW.The TF measurement was in accordance with the method used by Sanchez-Rangel et al. . The sup3 solution (w/v), 2 mL 5% H2SO4 (v/v) and 4 mL 5% H8MoN2O4 (v/v). After 15 min, the absorbance was taken at 705 nm by the UV-spectrophotometer.Vitamin C (VC) content was analyzed according to Li et al. . Fresh lVitamin A (VA) contents were determined using the plant vitamin A ELISA Kit , according to the manufacturer’s protocol. The absorbance was measured at 450 nm by the microtiter plate reader and the VA contents were recorded by comparing the absorbance to the standard curve.The overall data were firstly normalized using the method of Z-score and centered to the median of the entire sample set to lower the relatively large differences in different parameters. Then, the transformed data were visualized into heat map by TBtools software (v1.09867) using the function called Heatmap Illustrator .www.graphpad.com).Data were analyzed by a one-way analysis of variance (ANOVA) and two-way analysis of variance, SPSS software  with Tukey’s honest significant difference tests. All values were reported as the means of four replicates with standard deviations (SD). The principal component analysis was performed with GraphPad Prism 9  (p < 0.001), root dry weight (p < 0.05) and root to shoot ratio (p < 0.001)  . Under t< 0.001) . Both “Y< 0.001) . However< 0.001) . Meanwhi< 0.001) . Besides< 0.001) . The int< 0.001) .p < 0.001) (p < 0.001)  of lettuce leaf were significantly affected by light quality, cultivars and their interaction (< 0.001) . Lettuce< 0.001) . While b< 0.001) . In “Yanp < 0.001) (The cultivar differences showed that “Yanzhi” possessed better antioxidant capability (< 0.001) . Howeverp < 0.001) (p < 0.001). WB was effective in increasing the contents of TPC (22.62% and 39.46%) and TF (46.67% and 33.48%) in “Yanzhi” and “Red Butter”, compared to those under W. The TPC content of “Red Butter” was 43.05% lower while the TF content was 33.48% higher under WFR than W, respectively. The VC content of “Yanzhi” increased by 14.43% under WB, while that of “Red Butter” reduced by 23.26% by WFR  . Meanwhi% by WFR . Interesp < 0.001) (p < 0.05 or 0.001). In “Yanzhi”, the contents of SP and SS were not affected by light quality, while in “Red Butter” those were 20.06% and 47.21% higher in WFR than W, respectively. The SP content of “Red Butter” decreased by 17.04% under WB, compared to W. Both in “Yanzhi” and “Red Butter”, the nitrate content significantly increased, 11.86% and 9.97% under WFR, respectively.The nutritional compounds of different lettuce cultivars showed significant differences that “Yanzhi” possessed higher contents of SS and SP, while “Red Butter” had higher nitrate content (< 0.001) . The ligTo explore the response pattern of lettuce under different light quality and to statistically analyze the correlation among plant biomass, color parameters and phytochemicals, the principal compound analysis  and the From the whole, the first four principal components (PCs) of “Yanzhi” were connected to eigenvalues higher than 1 b and expAdditionally, the clustered heat maps well complemented and verified the results generated by PCAs. In “Yanzhi”, the light quality was divided into three main clusters a. The clCryptanthus, Guzmania and Hypoestes [Two cultivars of red-leaf lettuce were subjected to three light qualities in the PFAL; the cultivar responded differently to WFR and WB. The leaf color can be largely affected by environmental factors, especially the light conditions. Light spectra with higher percentage of blue light have been proven to positively impact on the accumulation of chlorophylls in lettuce, spinach, kale, basil pepper and cucumber leaves ,32,33, aypoestes . In contypoestes ,29,35. S2 and suppressing photorespiration [The blue light could also enhance the photosynthesis efficiency by regulating the stomatal opening, obtaining more COpiration . Consequpiration ,38. HoweWhile far-red light was reported to induce shade-avoidance syndromes and reduce the plant weight ,16, it f−1 FW) in the two cultivars were far below the general level (2.5~4.5 mg·g−1 FW) of lettuce [Furthermore, the heat map analysis showed that the nitrates and the fresh weight and dry weight of the edible parts in both lettuce cultivars were in the same cluster under different light treatments, indicating the importance of nitrogen to plant growth . WFR led lettuce . Thus, tIn general, blue light can strengthen the antioxidant capacities of vegetables through light signaling or creating light stress ,48,49. HPAL, F3H, CHS, ANS and GST [Light spectra also influence the phytochemical accumulation in plants. With respect to health-promoting compounds, TA, TPC and TF were determined in both lettuce cultivars . These m and GST ,59. ThusThe VC is known for its antioxidant properties. The VC functions as a redox buffer that reduces, and thereby neutralizes, reactive oxygen species . No chanSimilarly, the VA also has nutritional and antioxidant functions. The VA protects the cholesterol from oxidation by quenching oxidants like superoxide, hydroxyl and peroxide radicals . The VA In addition, the nutritional compounds such as SP and SS of two lettuce cultivars seemed to be less sensitive to WB, while these were significantly increased by WFR in both cultivars . These r−2·s−1 PPFD LED, the supplementary blue light could be applied to deepen the red color of lettuce leaves via accumulating the TA contents. Meanwhile, it benefited the flavor formation of lettuce by increasing the contents of TPC, TF and SS. Besides, the supplementary blue light drastically increased the VA content in two lettuce cultivars and the VC content in lettuce “Yanzhi”, which indicated that blue LEDs could be used to elevate the lettuce quality by making richer its health-promoting compounds. As for supplementary far-red light, it obviously favored the biomass of two lettuce cultivars through higher N uptake, which was of great significance to increase the vegetable production. In addition, although the far-red light was not as superior as blue light in improving crop qualities, it still had the ability to increase the contents of SP and SS in lettuce “Red Butter” and the VA content in the two lettuce cultivars. Thus, this study suggested that both far-red and blue LEDs have promising application prospects in PFALs as supplemental lights.In PFALs, under 250 μmol·m


The occurrence of hypoglycemia was affected by gender and nondiabetic patients (p < .001). In nondiabetic patients, hypoglycemia was affected by age (p < .05). For each year of age increase, the risk of hypoglycemia was reduced by 2.6%; the risk of hypoglycemia decreased by 3.7% when the waist circumference increased by 1 cm; the risk of hypoglycemia increased by 30.3% with each unit of TG increased; the risk of hypoglycemia decreased by 66.1% for each unit increased by TC; the risk of hypoglycemia increased by 1.715 times for each increased unit of HDLC; the risk of hypoglycemia increased by 1.185 times for each unit of LDLC. Women and nondiabetic people are more likely to have hypoglycemia. Among the group without diabetes, diastolic blood pressure, triglyceride, HDLC, and LDLC are risk factors of hypoglycemia.To further clarify the correlation between glucose and other biochemical indicators, to search the risk factors of hypoglycemia through a cross‐sectional study. Data were obtained from subjects who underwent health examination in the Health Promotion Center from July 2018 to June 2019. Hypoglycemia is defined as fasting blood glucose less than 4.2 mmol/L. All data were analyzed using Windows R software. A total of 23,935 subjects were included. There were significant differences in age, BMI, waist circumference, blood pressure, glucose, lipid, thyroid function, liver function, and kidney function between men and women ( To further clarify the correlation between glucose and other biochemical indicators, to search the risk factors of hypoglycemia through cross‐sectional study. Women and nondiabetic people are more likely to have hypoglycemia. Among the group without diabetes, diastolic blood pressure, triglyceride, HDLC, and LDLC are risk factors of hypoglycemia. Systolic blood pressure and diastolic blood pressure were measured after 15 min of rest. Serum fasting blood glucose (FBG), high‐density lipoprotein cholesterol (HDL‐C), low‐density lipoprotein cholesterol (LDL‐C), total cholesterol (TC), triglyceride(TG), uric acid(UA), aminotransferase(ALT), aspartate aminotransferase (AST), blood urea nitrogen(BUN), and creatinine(Cr) were measured using a Hitachi 7600 clinical analyzer . Free triiodothyronine (FT3), free thyroxine (FT4), thyroid‐stimulating hormone (TSH), thyroid peroxidase antibody (TPOAb), and thyroid globulin antibody (TgAb) were quantified using chemiluminescent enzyme immunoassays .Clinical information was completed by trained general practitioners. Waist circumference, height,and body weight were measured by a well‐trained nurse. Body mass index (BMI) was calculated as body weight (kg)/height squared . Chi‐square test was used to compare categorical variables, 33.1p < .05). Among them, the age, BMI, waist circumference, blood pressure, fasting blood glucose, glycosylated hemoglobin, blood lipid (except HDLC), and liver function index of male are higher than that of female; male's uric acid is higher than female's; male's thyroxine is higher than female's; the thyroid‐stimulating hormone of female is higher than that of male; the thyroid antibody of female is higher than that of male , and the possibility of hypoglycemia (<4.2 mmol/L) in women was significantly higher than that in men. (2) Hypoglycemia was affected by diabetic and nondiabetic patients (p < .001). The possibility of hypoglycemia (<4.2 mmol/L) in diabetic patients was significantly lower than that in nondiabetic patients (Table We found that: (1) The incidence of hypoglycemia was affected by gender (ts Table .3.4p > .05), indicating that the possibility of hypoglycemia (<4.2 mmol/L) was similar in different age groups. (2) In nondiabetic patients, hypoglycemia was affected by age (p < .05), suggesting that the possibility of hypoglycemia (<4.2 mmol/L) was different in different age groups  In diabetic patients, hypoglycemia was not affected by age  For each year of age increase, the risk of hypoglycemia was reduced by 2.6%; (2) the risk of hypoglycemia decreased by 3.7% when the waist circumference increased by 1 cm; (3) the risk of hypoglycemia increased by 30.3% with each unit (mg/dl) of TG increased; (4) the risk of hypoglycemia decreased by 66.1% for each unit (mg/dl) increased by TC; (5) the risk of hypoglycemia increased by 1.715 times for each unit increased unit (mg/dl) of HDLC; (6) the risk of hypoglycemia increased by 1.185 times for each unit (mg/dl) of LDLC.4Abnormal blood glucose is common in the clinic. Most hypoglycemic events are thought to be related to diabetes. However, hypoglycemia often occurs in nondiabetic patients. The occurrence of diabetes in nondiabetic inpatients is related to malnutrition, heart failure, malignant tumor, kidney disease, etc., and it is considered that the occurrence of such hypoglycemia is related to high mortality ; Writing‐original draft ; Writing‐review & editing . yue zhu: Data curation ; Formal analysis . liying chen: Supervision .


Aortic valve stenosis (AS) has a high prevalence and poor prognosis in patients who receive maintenance dialysis. However, few large-scale observational studies in Japan have investigated patients with AS who underwent dialysis. In this study, we investigated the prevalence and factors associated with AS in Japanese patients who underwent dialysis.2 were categorized into the AS group (G1). Patients with Vmax ≥ 3.0 m/s, mean PG ≥ 20 mmHg, or AVA ≤ 1.0 cm2 were categorized into the moderate and severe AS groups (G2). We performed multivariate logistic regression analysis and compared G1 and G2 with the non-AS group to determine the risk factors for AS. We also investigated the risk factors for aortic valve calcification, which is a pre-stage for AS.In this cross-sectional analysis, we enrolled patients who underwent dialysis and transthoracic echocardiography between July 1, 2017 and June 30, 2018. Patients with a maximum aortic jet velocity (Vmax) ≥ 2.0 m/s, pressure gradient (PG) between the left ventricle and ascending aorta (mean PG) ≥ 20 mmHg, or aortic valve area (AVA) ≤ 1.0 cmp < 0.05). These factors were converted into ordinal categories, and a multivariate logistic regression analysis was performed. Patients with serum phosphorus levels measuring 5.0–5.9 mg/dL and > 6.0 mg/dL showed a higher risk of AS than those with serum phosphorus levels measuring < 4.0 mg/dL . Aortic valve calcification was associated with age, long-term dialysis, diabetes mellitus, administration of vitamin D receptor activators, elevated serum calcium levels, and anemia .Of the 2,786 patients investigated, 555 (20.0%) and 193 (6.9%) were categorized into G1 and G2, respectively. Multivariate logistic regression analysis revealed that age, long-term dialysis, and elevated serum phosphorus levels were associated with AS in both the groups (Patients on dialysis showed a high prevalence of AS, which was associated with age, long-term dialysis, and elevated serum phosphorus levels.UMIN000026756, registered on March 29, 2017.The online version contains supplementary material available at 10.1186/s12882-022-02758-y. Chronic kidney disease (CKD) is a significant risk factor for cardiovascular disease (CVD) –5. The pAge-induced changes in the aortic valve, leading to sclerosis and calcification,  have repThe prevalence of hypertension, diabetes mellitus, and mineral and bone disorders (MBD) is higher in patients on dialysis compared to the general population. These conditions are associated with arteriosclerosis and CVD –28. The The study included patients aged ≥ 20 years who underwent outpatient maintenance dialysis for > 1 year across 58 hospitals in the Tokai region of Japan. The baseline was selected as the time of transthoracic echocardiography (TTE) performed between July 1, 2017 and June 30, 2018. Patients with a history of aortic valve surgery and those who refused to participate in the study were excluded. 2,916 patients were enrolled and the following data was recorded: Patients’ background, history, comorbidities, medications, laboratory data, chest radiography findings, electrocardiogram tracings, and TTE images. Out of the 2,916 patients enrolled in the study, 130 patients who met the following criteria were excluded: duration of dialysis < 1 year, unknown initiation of dialysis, history of aortic valve surgery, or TTE performed outside the observation period.2. Patients who met any of the criteria were categorized into the AS group (G1); the rest were categorized into the non-AS group. Among the patients in G1, the moderate and severe AS groups were categorized as G2, with the Vmax criterion changed from ≥ 2.0 m/s to ≥ 3.0 m/s without any change in the mean PG and AVA criteria. Fig. All the patients underwent a TTE to measure the parameters associated with AS and to evaluate for aortic valve calcification. The maximum aortic jet velocity (Vmax), pressure gradient (PG) between the left ventricle and ascending aorta (mean PG), and the AVA was recorded to evaluate for AS. Aortic stenosis was defined based on the 2014 American Heart Association/American College of Cardiology Guidelines for the Management of Patients with Valvular Heart Disease. It was defined as (a) Vmax ≥ 2.0 m/s, (b) mean PG ≥ 20 mmHg, or (c) AVA ≤ 1.0 cmBaseline data is presented as means (standard deviation), medians (interquartile range), or percentages for categorical measures. Fisher's exact test for nominal variables and the Mann–Whitney U test for continuous variables was used to compare the baseline data between G1 and the non-AS group and between G2 and the non-AS group. Factors associated with AS on univariate analysis were subjected to multivariate logistic regression analysis along with the factors associated with MBD. The following 14 variables were subjected to multivariate logistic regression analysis: sex, age, duration of dialysis, nephrosclerosis as the primary contributor to CKD, diabetes mellitus as a comorbidity, calcimimetic use, vitamin D receptor activator (VDRA) administration, phosphate binder use, serum albumin level, serum corrected calcium level, serum phosphorus level, serum intact parathyroid hormone level, serum C-reactive protein level, and serum hemoglobin level. Multivariate logistic regression analysis was performed with these 14 variables, and age, dialysis duration, and serum phosphorus levels that showed significant differences were converted into ordinal categories. Patients were categorized on the basis of age into the following groups: < 60, 60–69, 70–79, and ≥ 80 years. Multivariate logistic regression analysis was performed on these age ordinal categories and 13 other factors. Similarly, Patients were categorized into the following groups on the basis of the duration of dialysis and serum phosphorus levels: < 5, 5–9, 10–14, and ≥ 5 years and < 4.0, 4.0–4.9, 5.0–5.9, and ≥ 6.0 mg/dL, respectively.Patients were further categorized into those with and without aortic valve calcification (a pre-stage of AS), and a multivariate logistic regression analysis was performed. The Mann–Whitney U test was used to compare the Vmax, mean PG, and AVA between patients with and without aortic valve calcification.p < 0.05 was considered statistically significant.All statistical analyses were performed using the R software . All values were two-sided, and This study was performed in accordance with the Ethics of Clinical Research (The Helsinki Declaration) and was approved by the Fujita Health University Center for Clinical Trial and Research Support . All patients included in this study provided informed consent after the purpose of the study was explained to them. This study was registered in the Clinical Trial Registry  on April 1, 2017.2. Based on these findings, the moderate and severe AS groups were categorized as G2, with the Vmax criterion changed from ≥ 2.0 m/s to ≥ 3.0 m/s without any change in the mean PG and AVA criteria. Among patients in G1, 193 (6.9%) were categorized as G2. Out of the 193 patients, 98 (50.8%) showed Vmax ≥ 3.0 m/s, 107 (55.4%) showed mean PG ≥ 20 mmHg, and 102 (52.8%) showed AVA ≤ 1.0 cm2. Table In total, 2,786 patients were enrolled in the study; 555 patients (20.0%) were categorized into G1 and 2,231 (80.0%) into the non-AS group. Out of the 555 patients in G1, 514 (92.6%) showed Vmax ≥ 2.0 m/s, 107 (19.3%) showed mean PG ≥ 20 mmHg, and 102 (18.6%) showed AVA ≤ 1.0 cmp < 0.001; aOR 1.41, 95% CI 1.21–1.64, p < 0.001; aOR 1.16, 95% CI 1.06–1.28, p = 0.001, respectively). Comparison between G2 and the non-AS group showed similar tendencies in these three variables . In contrast, nephrosclerosis as a contributor to CKD was not associated with AS. Diabetes mellitus as a comorbidity was associated with AS when G1 was compared to the non-AS group ; however, no significant difference was observed between G2 and the non-AS group .We performed a multivariate logistic regression analysis to compare G1 and G2 with the non-AS group  than that of < 5 years of dialysis. The aOR increased with an increase in serum phosphorus levels. The aORs of the serum phosphorus levels in the 5.0–5.9 and > 6.0 mg/dL groups were 1.52 and 1.83-fold higher and 2.24 and 2.66-fold higher in G1 and G2, respectively, compared to serum phosphorus levels < 4.0 mg/dL.The aOR for AS was determined on the basis of age, duration of dialysis, and serum phosphorus levels  Patients with a history of aortic valve surgery were excluded; therefore, a serious risk factor associated with AS may have been disregarded. (ii) This study was a cross-sectional analysis; therefore, the results cannot demonstrate causality. (iii) The accuracy of TTE was unlikely to be consistent across patients in this multicenter study. Aortic valve calcification was based on the subjective interpretation of echocardiography findings by technicians at each facility. (iv) Data were not collected on whether the patients had bicuspid aortic valves, which is one of the major causes of aortic stenosis. This may have affected the results of this study. (v) The percentages of missing values for Vmax, mean PG, and AVA were 12.2%, 25.4%, and 20.9%, respectively. Although one or more of these factors were measured in all patients, missing data may potentially lead to misclassification of the AS groups. (vi) The multivariate logistic regression analysis was performed by complete cases analysis. This may have affected the results.Patients who received maintenance dialysis showed a high prevalence of AS, which was associated with age, hyperphosphatemia, and duration of dialysis.Additional file 1.


Dairy cows with high NUE , medium NUE , and low NUE  were selected from 284 Holstein dairy cows with the same diet. Measurement of the rumen fermentation indices showed that the proportion of propionate was higher in HE_HP cows and ME_MP cows than in LE_LP cows (P < 0.05). The diversity of rumen bacterial community was higher in LE_LP cows than in ME_MP cows and HE_HP cows by 16S rRNA sequencing analysis (P < 0.05). Moreover, at the genus level, the relative abundances of Succinivibrionaceae_UCG_001, uncultured_Selenomonadaceae, and Acidaminococcus were higher in HE_HP cows than in LE_LP cows (P < 0.05). Interestingly, we found that these bacteria were positively correlated with milk protein yield and negatively correlated with Δ15N (P < 0.05). However, Clostridia_UCG_014, Saccharofermentans, Bacilli_RF39, and Desulfovibrio were lower in HE_HP cows and ME_MP cows than in LE_LP cows (P < 0.05), which were negatively correlated with milk protein yield and positively correlated with Δ15N (P < 0.05). In conclusion, the study showed that the diversity and relative abundances of rumen bacteria differed among different NUE cows, indicating that rumen bacteriome contributes to nitrogen metabolism in dairy cows.High nitrogen utilization efficiency (NUE) is important for increasing milk protein production and decreasing the feed nitrogen cost and nitrogen emission to the environment. Currently, there is a limited whole picture of the relationship between ruminal bacteriome and the NUE of dairy cows, even though some information has been revealed about the bacteriome and milk or milk protein production of dairy cows. The purpose of this study was to compare the rumen bacterial community in dairy cows with different nitrogen utilization efficiency under the same diet. The natural abundance of  NUE can be affected by heredity, nutrition, and management , can be used to characterize the nutrition status of animals  can be especially used as a predictor to effectively assess NUE  cows than in low-MPY cows. Succinivibrio and Clostridium were enriched in low-MPY cows compared to high-MPY cows. Although specific bacteria were related to milk composition (Rumen microbiota can convert fiber into digestible substances such as volatile fatty acid (VFAs) and microbial proteins. Ruminal VFAs produced by rumen microbial fermentation can provide 70% of the energy requirements in dairy cows . Moreoveposition , there a15N and MPY, as NUE indicators, were used to discuss NUE’s relationship with the rumen bacteriome. The objective of the study was to reveal the potential effects of rumen bacteriome on NUE in dairy cows.In this study, Δg for 5 min to obtain plasma and stored at −20°C. The plasma 15N and diet 15N were determined by Elemental Analysis-Isotope Ratio Mass . Δ15N was calculated using δ15N plasma - δ15N diet  = 59 ± 9] dairy cows, 7 medium NUE  dairy cows, and 7 low NUE  dairy cows were selected . A total of 284 healthy lactating Holstein dairy cows  were selected from 4,086 dairy cows in a commercial dairy farm. The dairy cows were fed an identical diet  and had 15N diet . Based oselected . The rumg at 4°C for 5 min to obtain the supernatant. Then, 600 μl of the supernatant was mixed with 50 μl of 25% metaphosphoric acid and was centrifuged at 12,000 × g for 20 min to obtain the supernatant for the analysis of ammonia nitrogen and VFAs. The ammonia nitrogen concentrations were determined using phenol-sodium hypochlorite colorimetric analysis . After the library quality control was finished, the library was sequenced on an Illumina HiSeq platform .Total DNA of rumen liquid was extracted from the rumen liquid of 21 dairy cows using cetyl trimethyl ammonium bromide  plus bead-beating method . The qua15N was generated using RevolverMaps.1Reads were removed primers, demultiplexed, and quality-filtered using Vsearch . The rea15N, rumen fermentation parameter concentrations , and alpha diversity indices of HE_HP cows, ME_MP cows, and LE_LP cows were analyzed using one-way ANOVA. The relative abundances of bacteria and alpha diversity indices among the three groups were compared using Kruskal–Wallis test. Dunn’s multiple-comparisons test was used for multiple comparisons of means among the three groups. Statistical analysis was performed using GraphPad Prism 7.0, with P < 0.05 considered as statistical significance. LEfSe was used to compare the relative abundances of rumen bacteria in HE_HP cows, ME_MP cows, and LE_LP cows, and significant differences were considered by a LDA score >2 and P < 0.05. Correlation analysis between the specific bacteria and NUE was performed using Spearman’s correlation.Lactation performance , Δ15N was 2.30 ± 0.05 ‰, the composition of acetate was 60.22 ± 0.60%, propionate was 27.15 ± 0.57%, isobutyrate was 0.47 ± 0.02%, butyrate was 9.79 ± 0.22%, isovalerate was 0.84 ± 0.04%, valerate was 1.52 ± 0.08%, and NH3–N was 8.85 ± 0.67 mg/dl. Milk yield and MPY in HE_HP were the highest among the three groups (P < 0.01) and in LE_LP cows were the lowest. In contrast, Δ15N in HE_HP cows was the lowest and in LE_LP cows was the highest . Furthermore, the ruminal levels of NH3–N and the proportion of butyrate and valerate were not different among the three groups (P > 0.05). However, the HE_HP cows exhibited a significantly lower concentration of acetate (58.89%) and a higher concentration of propionate (28.87%) compared to LE_LP cows (P < 0.05).As to the selected 21 dairy cows, the milk yield was 44.37 ± 1.75 kg/day (mean ± SEM), the MPY of 21 dairy cows was 1.50 ± 0.06 kg/day, ΔP > 0.05) were not significantly different in HE_HP cows, ME_MP cows, and LE_LP cows (P < 0.01) were significantly higher in LE_LP cows compared to HE_HP cows and ME_MP cows (P < 0.01), while the ME_MP group had no significant difference compared to the HE_HP group , Succinivibrionaceae_UCG_001 (23.42 ± 16.38%), Muribaculaceae (4.61 ± 3.95%), and Succiniclasticum (3.35 ± 2.91%) were the most abundant genera , Firmicutes (30.25 ± 11.96%), and then Proteobacteria (25.78 ± 14.97%) . At the t genera .Succinivibrionaceae_UCG_001, uncultured_Selenomonadaceae, and Acidaminococcus was higher in HE_HP cows (P < 0.05). Moreover, Clostridia_UCG_014, Saccharofermentans, Bacilli_ RF39, and Desulfovibrio were higher in LE_LP cows (P < 0.01). There were differences in the abundances of bacterial taxa among the three groups , and there was 20% less genus Succinivibrionaceae_UCG_001 in LE_LP cows than in the other two groups. Moreover, the most differently abundant OTUs belonged to the phylum of Firmicutes.The different abundant bacteria at the genus level among the HE_HP cows, ME_MP cows, and LE_LP cows were revealed by LEfSe . The rele groups . Meanwhie groups . The gen15N and MPY using Spearman’s correlation. The correlation analysis revealed that 32 bacterial taxa were significantly correlated with Δ15N and MPY . Among them, Succinivibrionaceae_UCG_001, uncultured_Selenomonadaceae, Acidaminococcus, Eubacterium_xylanophilum_group, and Lachnospiraceae_NC2004_group were positively correlated with MPY and negatively correlated with Δ15N. The scatter plots of the specific bacteria and Δ15N and MPY are shown in Clostridia_UCG_014, Saccharofermentans, Bacilli_RF39, and Desulfovibrio were negatively correlated with MPY and positively correlated with Δ15N (P < 0.05).In the study, in all of the 326 encountered genera, the relative abundance of 127 bacterial taxa was >0.01%. We analyzed the 127 bacterial taxa and Δ and MPY . The res15N .At present, the most effective way to improve the NUE of dairy cows is to match the dietary nitrogen content and energy supply during rumen fermentation by changing the diet composition . In addi15N . The Δ15 of cows . In receficiency . MPY is ficiency . HoweverThe rumen bacteria play an important role in dietary nutrient digestion and absorption. Furthermore, we detected that the diversity of rumen bacteria and specific rumen bacterial taxa contributed to NUE among the HE_HP cows, ME_MP cows, and LE_LP cows. The alpha diversity indices of Shannon and Simpson showed a lower bacterial diversity in HE_HP cows. Anaerobiospirillum, Succinivibrionaceae_UCG_001, Ruminobacter, Succinivibrionaceae_UCG_002, Succinimonas, and Succinivibrio), was the most active rumen bacterial family in beef . Thus, it might demonstrate that these four bacteria were negatively associated with NUE. Clostridia_UCG_014 is a kind of pro-inflammatory bacteria .MnL contributed to conceptualization, software, data curation, and writing—original draft. HZ performed data curation. MgL took charge of the software. NZ was in charge of supervision. SZ contributed to writing—review and editing. JW was in charge of administration. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Dear Editor,Statistical analysis. The authors mentioned “discrete variables were reported as numbers ”, but the statistical methods are not mentioned. A discrete variable is a variable whose value is obtained by counting. In contrast, continuous variable is a variable whose value is obtained by measuring [easuring . OperatiStudy population. In the study population, the authors mentioned that “with a minimum of 6 months follow-up” were included in the study. However, in the results, the authors also mentioned that “all patients received at least 1 year of clinical follow-up”; it seems to be a contradiction.Results. In the results, surgical time of FNS and CCS was 79.75 ± 26.35 min vs. 64.58 ± 18.56 ml, which is incorrect. It should be 79.75 ± 26.35 min vs. 64.58 ± 18.56 min.Recently, Hu et al. have published the article “Clinical outcome of femoral neck system versus cannulated compression screws for fixation of femoral neck fracture in younger patients” . We read


Artemisia argyi extract (AE) has excellent antibacterial and anti-inflammatory properties. In this research, we developed AE loaded composite hydrogel scaffold based on methacrylate gelatin (GelMA)/methacrylate hyaluronic acid (HAMA) and mesoporous silica nanoparticle (MSN) as sustained-release drug carrier vehicles for the treatment of chronic wounds. The presented GelMA/1%HAMA hydrogel possessed stable rheological properties, suitable mechanical properties, appropriate biodegradability, swelling, sustained-release AE capacity. In vitro antibacterial and cell experiments showed that the GelMA/HAMA/MSN@AE hydrogel had excellent antibacterial activity and biocompatibility and induced macrophages to differentiate into M2 phenotype. In vivo wound healing of rat full-thickness cutaneous wounds further demonstrated that the prepared GelMA/HAMA/MSN@AE hydrogel could significantly promote chronic wound healing by upregulating the expression of IL-4, TGF-β1, CD31, and α-SMA but downregulating the expression of TNF-α and IFN-γ and promoting M1-M2 macrophages polarization. Altogether, we believe that the GelMA/HAMA/MSN@AE hydrogel will have wide application prospects in healing chronic wounds.Chronic wounds are a major health problem with increasing global prevalence, which endangers the physical and mental health of those affected and is a heavy burden to healthcare providers.  Chronic wounds have been regarded as a “silent epidemic,” presenting a major challenge for the healthcare system , among wThe impact of chronic nonhealing wounds is enormous, resulting in prolongation of hospitalization and economic burden on patients with the use of expensive wound care products, which further increases the burden on the national healthcare system . MeanwhiArtemisia argyi has been widely used in promoting wound healing. The active compounds from Artemisia argyi leaves can easily permeate the skin and have the effects of activating blood circulation, transforming qi, dispelling dampness, dissipating cold, eliminating swelling, resolving blood stasis, relieving pain, and promoting necrotic tissue shedding (Artemisia argyi extract (AE) has a variety of pharmacological activities, such as anti-allergic, anticoagulant, complement activation, anti-inflammatory, antibacterial, antiviral, and sedation, which can reduce tissue edema, promote granulation growth for wound tissue, and avoid adverse consequences of long-term antibiotic treatment  light condition. The porous structure, physical properties, drug release, and antibacterial properties of this hydrogel were characterized. The slow and sustained release of AE in the hydrogel can be realized by a large specific surface area and large pore volume of MSN. In addition, the cytocompatibility in hydrogels was studied in detail by CCK-8 staining and living/dead cell staining. Moreover, an excellent therapeutic effect for promoting the repair of skin defects was shown in Artemisia argyi extract (AE) was purchased from Anhui Chinature Biological Co., Ltd. . Gelatin  was obtained from the Sigma-Aldrich Chemical Company . Hyaluronic acid  and methacrylic anhydride (MA) were purchased from Macklin Biochemical Technology Co., Ltd. . Phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) was obtained from Yinchang New Material Co., Ltd. . Live/dead cell staining kits and Cell Counting Kit-8 (CCK-8) were obtained from BestBio Co., Ltd. . The bacteria strains of Escherichia coli  and Staphylococcus aureus  were obtained from Luwei microbial Sci&Tech Co., Ltd. . Dulbecco’s Modified Eagle Medium (DMEM) and Fetal Bovine Serum (FBS) were obtained from Gibco .GelMA was prepared according to the previous method . BrieflyHAMA was prepared as previously described with slight modification . BrieflyUniform-sized MSN was synthesized according to a previously reported method . TotallyIn order to prepare AE-loaded MSN (MSN@AE), 10 mg of AE and 50 mg MSN were added into 5 ml of 25% ethanol solution and stirred for 24 h at 37°C. The mixture was centrifuged, and the AE concentrations of residues in the supernatant were measured by a UV spectrophotometer to determine the MSN loading capacity .The freeze-dried GelMA and HAMA were dissolved in PBS at 60°C to make the final GelMA concentrations of 10% (w/v) with different HAMA concentrations of 0.5% (w/v), 1% (w/v), and 2% (w/v), named GelMA, GelMA/0.5%HAMA, GelMA/1%HAMA, and GelMA/2%HAMA, respectively. Then, 0.1% (w/v) photo-initiator LAP was added, and the prepolymer solution then exposed to the UV light (365 nm) for 30 s. For the preparation of GelMA/HAMA/MSN and GelMA/HAMA/MSN@AE, the MSN 0.5% (w/v) and MSN@AE 0.5% (w/v) were added indirectly into above GelMA10% (w/v)/HAMA 2% (w/v) mixed solutions, respectively.1H NMR using a nuclear magnetic resonance spectrometer . Fourier transform infrared spectroscopy (FTIR) spectra of Gel, GelMA, MSN, MSN/AE, GelMA/HAMA/MSN, and GelMA/HAMA/MSN@AE were measured by Fourier transform infrared spectrometer .The chemical structure of GelMA and HAMA were characterized by The surface area, pore size, and pore volume of MSN and MSN/AE were measured by the BET method using Micromeritics ASAP2460 instrument . The micromorphology of the composite hydrogels was analyzed by scanning electron microscopy . The average pore size of hydrogels was calculated by Nanomeasure software. Thermal gravimetric analysis (TGA) measurements of MSN, MSN/AE, GelMA/HAMA/MSN, and GelMA/HAMA/MSN@AE were performed with a TGA thermogravimetric analyzer .t and W0 mean the weight of hydrogel at times t and 0, respectively.The swelling ratio of the composite hydrogel was investigated using a gravimetric method . The tes1) and the mass of the dry sample immersed in anhydrous ethanol (density = ρ) for 2 h (W2) were weighed. The porosity (P) was calculated according to the following formula:The hydrogel was soaked in PBS for 24 h to reach swelling equilibrium and then freeze-dried. The volume (V) of the freeze-dried hydrogel was accurately measured with a Vernier caliper. The mass of the dry sample . The change curves of the storage modulus (G′) and loss modulus (G″) of the hydrogels were detected. In order to assess the shear viscosity behavior, the flow sweep assay was tested with a shear frequency range of 0.1–10 Hz at room temperature. Moreover, the viscoelastic region with a fixed frequency of 1 Hz was recorded over time performed at a strain of 1%.The compression modulus of GelMA/HAMA hydrogels was recorded by a universal testing machine . The 500 μL hydrogels were prepared as a cylindrical shape (height = 6 mm and diameter = 12 mm). The compressive strain rate was fixed at 0.05 mm/s, and the strain level reached 60% of the maximum. Meanwhile, Young’s modulus could be calculated as the slope at the initial linear region.t and W0 corresponded to the weight of lyophilized hydrogel at times t and 0, respectively.The 300 μL hydrogel samples were immersed in a PBS solution containing either 0 or 1000 U/ml lysozyme at 37°C. At given time points, the samples were taken out from the solution and rinsed three times with ultra-pure water. The dry weight of samples was weighed after freeze-drying. The weight loss ratio was calculated according to the following equation:The composite hydrogels were immersed in 10 ml PBS and incubated at 100 rpm at 37°C. At given time points, 1 ml supernatant was collected and replaced with 1 ml fresh PBS. The AE concentration was measured using a UV spectrophotometer at λ = 345 nm. The concentration of AE released from hydrogels was back-calculated using a standard curve.E. coli) and Gram-positive (S. aureus) were cultured in a sterilized LB liquid medium and diluted to an optical density of 600 nm of 0.1. Then, 400 μL tested hydrogels were prepared in a 48-well plate. Each sample was cocultured with 1.8 ml bacterial suspension and incubated at 37°C for 4 h under constant shaking. The resulting bacterial liquid (100 μL) was plated on an LB agar plate after a series of dilutions and incubator for 24 h. In order to determine the antibacterial activity, the antibacterial rate was determined according to the equation as follows:control and Nsample are the numbers of bacterial colonies of the GelMA/HAMA hydrogel sample and the hydrogel with different concentrations of AE, respectively.Gram-negative (2.Mouse fibroblast cells (L929 cells) were cultured in DMEM supplemented with 10% FBS, 100 μg/ml streptomycin, and 100 U/ml penicillin at 37°C in an incubator containing 5% CO4 cells/ml) was added to the hydrogel surface. After incubation for 1, 2, and 3 days, each well was immersed in 300 µL CCK-8 and incubated in a 5% CO2 humidified incubator at 37°C for 1 h. The absorbance of each well was measured at 450 nm using a microplate reader .The CCK-8 and live/dead cell staining methods were used to assess the cytotoxicity of hydrogels. Briefly, 500 μL hydrogels were prepared in 48-well plates and irradiated by UV light for 30 s. Then, 400 μL cell suspension . Meanwhile, the tested hydrogels were fixed to evaluate L929 cells morphology. TRITC Phalloidin was used to counterstain cytoskeleton, and diamidino-2-phenylindole (DAPI) was used to counterstain cell nuclei.4 cells/well and cultivated overnight at 37°C in an incubator containing 5% CO2 for later use. Secondly, the sterilized hydrogel was immersed in the DMEM (containing LPS); Then, the hydrogel extract was collected after culturing at 37°C for 24 h and cocultured with cells for 48 h. The sample without the hydrogel extract was used as the control group. Characterization of M1/M2-polarized macrophages was determined by the WB technique. Raw 264.7 macrophages were lysed using RIPA lysis buffer supplemented with 1% PMSF on ice for 20 min and subjected to Western blot analysis. Each protein sample was subjected to electrophorese by 10% sodium dodecyl sulphate-polyacrylamide gel (SDS-PAGE). Then, the protein was transferred to polyvinylidene difluoride (PVDF) membrane, further incubated with different primary antibodies, anti-rabbit iNOS , and anti-rabbit CD206 , overnight at 4°C and treated with horseradish peroxidase- (HRP-) conjugated secondary antibodies for 1 h at 22°C. Blots were developed using an enhanced chemiluminescent reagent , and the signals were detected via X-ray films. The expression levels of protein were quantified by IPP 6.0 software.The RAW 264.7 cells were seeded in a 6-well plate containing DMEM at densities of 1 × 10via intraperitoneal injection (45–60 mg/kg) prior to surgery. The backs of rats were depilated with a depilatory cream and disinfected with iodophor. Four 12 mm diameter circular full-thickness skin injuries were created on the dorsum of rats by excising the dorsum with a distance of 2 cm. Then, the wound was coated with gauze, GelMA/HAMA hydrogel (200 μL), GelMA/HAMA/MSN@AE hydrogel (200 μL), commercially available hydrocolloid dressing, respectively. The hydrogels were fixed by Tegaderm (3M) to cover and protect the wound area. Sterile medical cotton gauze was served as the control group. Following surgery, each rat was housed in a single cage, and the wound hydrogels were replaced every 3 days.All animal experimental protocols have been reviewed and approved by the Animal Protection and Use Committee of Jinan University. Sprague Dawley female rats, 200–250 g, were used in this study. Rats were anesthetized by 3% pentobarbital The physical appearance was photographed, and the area of wounds was calculated using the IPP 6.0 software on days 3, 7, 10, and 14 after surgery. The wound closure rate was expressed as the percentage values of the day 0 measurements. The wound tissues, including the wound site and unwounded area, were collected for the following experiment.Each wound tissue was isolated and fixed in 4% paraformaldehyde solution for 24 h, then, dehydrated through a graded series of ethanol, transferred into dimethylbenzene, and embedded in paraffin. Serial sections (4 μm thick) were cut and stained with hematoxylin and eosin (H&E) and Masson’s trichrome staining. Images of the stained sections were captured by the microtome .1 , tumor necrosis factor-α , interleukin-4 , interferon-γ , iNOS , and CD206 , were added at 4°C overnight. Next, these sections were incubated with HRP-labeled goat anti-rabbit IgG secondary antibody for 50 min at room temperature. Subsequently, the sections were reacted with DAB solution after being washed in PBS, and the nuclei were counterstained with DAPI. Images of smear specimens were also collected by the inverted fluorescence microscope.After paraffin sections were rehydrated, slices were incubated in an antigen retrieval solution and blocking serum. Then, the primary antibodies, α-SMA , CD31 , transforming growth factor-βp < 0.05 was defined as statistically significant: *p < 0.05, **p < 0.01, and ***p < 0.001.All quantitative experimental values from the studies were presented as means ± standard deviation. Analysis of variance  was used to determine significant differences between two groups. A value of We have developed a UV-cross-linked biocompatible hydrogel composed of photosensitive GelMA and HAMA. GelMA and HAMA were combined by cross-linking to improve mechanical properties and biological stability. Moreover, AE was used as an anti-inflammatory, antimicrobial agent, which was encapsulated by MSN to improve its bioavailability and loaded into the GelMA/HAMA hydrogel for application in faster healing of chronic wounds. These hydrogels also exhibited good antibacterial properties and comparable modulus to human soft tissue .2/g, the adsorption cumulative pore volume was 1.488,417 cm³/g and 1.377,355 cm³/g, and the adsorption average pore width was 6.9999 and 6.9297 nm, respectively. N2 adsorption/desorption isotherms of MSN and MSN@AE are shown in TEM imaging of MSN showed a uniform spherical shape with average diameters of ∼70 nm . The BET1H-NMR. The distinctive double peaks (δ = 5.4 and 5.6 ppm) were observed in GelMA (δ = 5.7 and 6.1 ppm) were observed in HAMA . Bands were observed at 1,656, 1,539, and 1,448 cm−1 belonging to the CH2 wagging vibrations of amide I, plane bending of N−H bond of amide II, and C−H stretching vibrations of amide III, respectively. Meanwhile, the characteristic peak intensities of amide III, amide II, and C=O in GelMA had a partial shift and change compared with gelatin, which demonstrated the newly formed amide bond. As shown in −1 is related to the stretching vibration of the OH group. The strong absorption band at 1,083 cm−1 is the characteristic absorption band of Si-O-Si antisymmetric stretching vibration. However, there are no other absorption peaks related to C-H, which indicates that the organic guides in the MSN have been removed at high temperatures. Therefore, FTIR showed that the composition of the MSN should be silica. There was a large wide peak in 3,500–3,700 cm−1, representing the characteristic absorption peak of hydroxyl in the three materials.As shown in Artemisia argyi, the TG curves of MSN and MSN@AE were almost identical  hydrogels, which demonstrated the outstanding potential for antibacterial performance of GelMA/HAMA/MSN@AE. The excellent antibacterial properties of the GelMA/HAMA/MSN@AE hydrogel were attributed to the AE, which could increase the permeability of bacterial cell membranes and inhibit the synthesis of bacterial nucleic acid, resulting in killing bacteria. We believed that GelMA/HAMA/MSN@AE had excellent antibacterial properties through the sustained release of the AE decomposing from MSN@AE, which could effectively eliminate the risk of bacterial infections in skin wounds.To further evaluated the antibacterial activity of hydrogels fections . As showThe biocompatibility of composite hydrogel is a crucial factor for functional tissue regeneration and a standard for safe application in the biomedical field . The cytin vitro anti-inflammatory effect of the GelMA/HAMA/MSN@AE hydrogel. As shown in It has been recognized that the inflammatory microenvironment plays a critical role in regulating wound healing . MacrophTo further verify the clinical potential of hydrogel, a full-thickness cutaneous wound model in rats was applied to assess the wound repair capability of the GelMA/HAMA/MSN@AE hydrogel. As shown in On day 3, the wound size of the gauze, GelMA/HAMA hydrogel, GelMA/HAMA/MSN@AE hydrogel, and commercially available hydrocolloid were 96.1 ± 8.1%, 82.1 ± 8.0%, 69.3 ± 10.2%, and 77.4 ± 8.9%, respectively. After 7 days of operation, the wound size of the GelMA/HAMA/MSN@AE group was 25.7 ± 2.6%, which was significantly lower than that of the control group (44.6 ± 25.0%). On day 10, it can be observed that GelMA/HAMA/MSN@AE hydrogels displayed superiority in promoting wound healing and the wound was nearly closed, while the gauze group, GelMA/HAMA, and commercially available hydrocolloid group still had a wound area of 11.7 ± 0.8%, 9.9 ± 1.1%, and 9.2 ± 0.7%, respectively. In summary, GelMA/HAMA/MSN@AE hydrogels showed the smallest wound size, which was attributed to the antibacterial and anti-inflammatory properties of AE, gelatin, and hyaluronic acid’s desirable function in the promotion of wound healing, and the moist wound environment provided by the hydrogel dressing. AE could accelerate different wound healing phases , as it could effectively scavenge reactive oxygen species (ROS) and exert anti-oxidant effects during the inflammatory phase, promote fibroblast migration, collagen formation, and epithelialization in the proliferation phase, and increase the number of cytokines in the remodeling phase.in vivo. As shown in Wound tissue regeneration and re-epithelialization is one of the evaluation criteria in the process of wound healing . H&E staIn addition, collagen deposition plays a significant role in the remodeling period of wound healing and is beneficial to wound contraction. Masson staining was carried out to evaluate the collagen deposition in the regenerated skin tissue. As shown in Neovascularization is one of the key factors to promote wound healing, which reflects the degree of skin tissue regeneration and functional recovery . The mecInflammation will hinder the healing process of the wound bed, but anti-inflammatory properties of AE have been unequivocally established in previous studies . To furtAs shown in Meanwhile, macrophages can coordinate the complex processes of cell proliferation, infection, inflammation, and functional tissue regeneration in the wound , respondAdditionally, although anti-inflammatory M2 phenotype macrophages maker (CD206) was also detected in the GelMA/HAMA and commercially available hydrocolloid groups, the GelMA/HAMA/MSN@AE group showed the most expression of CD206. All results demonstrated that GelMA/HAMA/MSN@AE hydrogel could upregulate the expression of CD206 but downregulate the expression of iNOS, which promoted the transformation of macrophages from inflammatory phenotype to a reparative phenotype and induced the wound to enter the proliferative phase. Summary, all results indicated that compared to other groups, the GelMA/HAMA/MSN@AE hydrogel group exerted anti-inflammatory effects in the chronic wound through enhanced expression of CD31, α-SMA, IL-4, and TGF-β1, reduced the expression of TNF-α and IFN-γ, and facilitated the M1-to-M2 transition of macrophages in the early stage of wound healing, playing a beneficial role in wound closure.Artemisia argyi extract (AE) was successfully prepared. GelMA/HAMA/MSN@AE hydrogel has stable rheological properties, suitable mechanical properties, appropriate biodegradability, swelling ratio, and sustained-release AE capacity. AE could increase the permeability of bacterial cell membrane and inhibit the synthesis of bacterial nucleic acid, so GelMA/HAMA/MSN@AE hydrogel had excellent antibacterial activity. AE could regulate inflammation, and the synthetic materials of GelMA and HAMA are natural cytoplasmic matrix materials, so GelMA/HAMA/MSN@AE hydrogel had good biocompatibility and promoted M1-M2 transformation of macrophages. AE could promote the growth of collagen fibers after full-thickness skin trauma and inhibit inflammation. Hydrogel could protect the wound from external stimulation and provide a moist environment. GelMA/HAMA/MSN@AE hydrogel could increase collagen deposition, promote angiogenesis, and regulate inflammation in chronic wound healing of rats. Overall, our results concluded that GelMA/HAMA/MSN@AE hydrogel with the sustainable release ability of AE exhibited an excellent property in promoting chronic wound healing.The purpose of this work is to prepare hydrogels with antibacterial properties, immune regulation, anti-inflammation, and good biocompatibility for the treatment of wounds. Then, GelMA/HAMA hydrogel with MSN-loaded