ObjectiveTo investigate whether Bushen Huoxue prescription improves embryo implantation through regulating exosomal miRNA to enhance maternal-fetal interface communication based on Bushen Huoxue therapy. MethodsIn the animal experiment， all the rats （except for the blank group） were administered hydroxyurea （450 mg·kg^-1） via gavage for 10 d， as well as epinephrine （0.3 mg·kg^-1） and mifepristone （5.5 mg·kg^-1） via subcutaneous injection for 7 d to establish an implantation disorder model of kidney deficiency and blood stasis type. The Bushen Huoxue prescription （BSHX） groups were administered the prescription at different doses （7.30 g·kg^-1 for the high-dose group， 3.65 g·kg^-1 for the medium-dose group， and 1.83 g·kg^-1 for the low-dose group） via gavage. The dydrogesterone group was administered the corresponding medicine （2.63 mg·kg^-1） via gavage. After intervention for 10 days， uterine histopathological changes were observed via hematoxylin-eosin （HE） staining. Mucin （MUC1）， forkhead box protein O1 （FoxO1）， and homeobox A10 （HoxA10） expression levels were detected by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot. Cell experiment selected primary endometrial epithelial cells （EEC） and trophoblast cells （TC） as research subjects. Exosome-free medicated serum was prepared by ultracentrifugation and cultured in complete medium. Exosomes were isolated from cell supernatants by ultracentrifugation for cross-co-culture. After 48 h， migration and invasion abilities were assessed by scratch and Transwell assays. Sequencing was then performed on EEC-origin exosomal miRNA. ResultsThe model rats exhibited thin endometrium， along with reduced blood vessels， glandules， and pinopode numbers. BSHX improved endometrial morphology and increased pinopode numbers. MUC1， FoxO1， and HoxA10 expressions were downregulated in the model rats， while these parameters were upregulated after BSHX medium- and high-dose intervention. In the cell experiment， after exosome-free medicated serum intervention for 24 h， migration and invasion abilities were enhanced in the BSHX groups （P<0.01）. In EEC-origin exosomal miRNA sequencing， gene ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） analyses revealed enrichment in biological processes （gastrulation， neuronal differentiation， alongside cell development and regeneration）， involving the mitogen-activated protein kinase （MAPK）， FoxO1， Wnt， mammalian target of rapamycin （mTOR）， and tumor necrosis factor （TNF） signaling pathways. ConclusionBSHX promotes embryo implantation by improving endometrial receptivity via regulating exosomal miRNA. These findings provide potential targets for exosomal miRNA-based assisted reproductive strategies and a novel theoretical basis for infertility treatment by traditional Chinese medicine.

Renal lymphatic vessel density is clinically relevant in kidney disease but is still assessed by slow, subjective visual estimation. We evaluated a weakly supervised, attention-based multiple-instance learning framework for automated detection and quantification of renal lymphatic vessel density on D2-40-stained whole-slide images (WSIs). Methods: Two independent internal datasets from Tongji Hospital were collected, including 198 cases of chronic kidney disease (CKD) and 50 cases of hypertensive nephropathy (HTN). All biopsies were immunohistochemically stained for D2-40 and digitized as WSIs. Pathologists provided only slide-level labels (D2-40 high vs. D2-40 low). Tissue regions were automatically segmented, tiled into patches, and encoded using a pretrained convolutional neural network. Patch embeddings were then analyzed with a clustering-constrained attention multiple-instance learning (CLAM) model. Unlike conventional multiple-instance learning (MIL) methods that only weight instances, CLAM jointly performs attention-based instance selection and instance-level clustering to distinguish positive from negative evidence within each slide, yielding more discriminative slide-level features and interpretable attention maps. Performance was compared with a classic MIL model trained on the same features. Results: CLAM achieved area under the receiver operating characteristic curves of 0.942 and 0.858 on the CKD and HTN datasets, respectively, outperforming classic MIL (0.866 and 0.801). Attention maps highlighted lymphatic-rich regions consistent with renal pathologists’ assessments. Conclusions: This clustering-constrained, attention-based weakly supervised framework enables fully automated, reproducible quantification of renal lymphatic vessel density from WSIs, providing renal pathologists with rapid visual and numerical support for diagnosis and risk stratification in CKD and HTN.

Objective To investigate the clinical prognostic value of the prognostic nutritional index (PNI) in patients with renal cell carcinoma (RCC) undergoing partial nephrectomy.Methods A retrospective analysis was conducted on the clinical data of RCC patients who underwent partial nephrectomy at our hospital during 2020 and 2024.The optimal cutoff value of PNI was determined based on the maximum Youden index from the receiver operating characteristic (ROC) curve.The clinicopathological characteristics, overall survival (OS), and progression-free survival (PFS) between patients with different PNI levels were compared.Cox regression analysis was used to explore the independent prognostic factors for RCC patients. Results Based on the ROC curve, the optimal cutoff value corresponding to the maximum Youden index was calculated to be 46.675, at which point the sensitivity was 65.3% and the specificity was 62.2%.Of the 815 patients included in the study, 293 (36.0%) patients had a PNI＜46.675.Cox regression analysis confirmed that preoperative PNI, tumor diameter, ISUP classification, and ECOG score were independent risk factors for postoperative OS in patients with RCC who underwent PN (P＜0.05).In addition, preoperative PNI, tumor diameter, ISUP classification and ECOG score were independent risk factors for postoperative PFS (P＜0.05).After covariates in the multivariable Cox regression model were adjusted, patients in the low PNI group demonstrated significantly shorter overall survival (HR=2.392, P＜0.001) and significantly poorer progression-free survival (HR=2.076, P＜0.001) compared to those in the high PNI group.Conclusion Preoperative PNI has significant clinical value in evaluating the prognosis of RCC patients undergoing partial nephrectomy.An elevated preoperative PNI is associated with a better prognosis.

ObjectiveTo investigate the mechanism by which Luoshi Neiyi prescription treats endometriosis （EMs） through the Toll-like receptor 4 （TLR4）/nuclear factor-κB （NF-κB） signaling pathway. MethodsAnimal experiments were conducted with 50 female SD rats， which were randomized into a sham operation group （10 rats） and a modeling group （40 rats）. An autologous endometrial transplantation method was used for the modeling of EMs. The 36 successfully modeled rats were randomly allocated into four groups （n=9 each）： The model group， the low-dose （7.87 g·kg^-1） Luoshi Neiyi prescription group， the high-dose （15.74 g·kg^-1） Luoshi Neiyi prescription group， and the dienogest （0.20 mg·kg^-1） group. The physiological status and ectopic lesion volumes of rats in each group were observed. Hematoxylin-eosin （HE） staining was used to observe the morphology of the eutopic endometrial tissue. Enzyme-linked immunosorbent assay （ELISA） was employed to measure the levels of inflammatory factors including interleukin-1β （IL-1β） and prostaglandin E₂ （PGE₂） in the serum of EMs rats. Immunohistochemistry was used to detect the expression of matrix metalloproteinase-9 （MMP-9） and vascular endothelial growth factor A （VEGFA） in the eutopic endometrial tissue. Western blot was adopted to determine the protein levels of TLR4， myeloid differentiation factor 88 （MyD88）， phosphorylated nuclear factor-κB （p-NF-κB）/NF-κB， MMP-9， and VEGFA in the eutopic endometrial tissue. In the cell experiments， the cell-counting kit-8 （CCK-8） assay was employed to screen the optimal concentration of Luoshi Neiyi prescription-containing serum， and a scratch assay was performed to assess the migration ability of iheESCs cells. Interventions with Luoshi Neiyi prescription-containing serum， resatorvid （TAK-242， a TLR4 inhibitor）， and lipopolysaccharides （LPS， a TLR4 agonist） were conducted， and Western blot was used to detect the expression of proteins related to the TLR4/NF-κB signaling pathway. ResultsThis experiment successfully replicated 36 EMs rat models. Compared with the sham operation group， the model group exhibited visible ectopic lesions on the abdominal wall and an increase in the writhing response score （P<0.01）. Furthermore， HE staining revealed the model group exhibited a thickened eutopic epithelium， stromal cell disarrangement， and evident infiltration of inflammatory cells. In addition， the model group showed elevated serum levels of IL-1β and PGE₂ （P<0.05， P<0.01）， increased positive expression of MMP-9 and VEGFA in the ectopic endometrial tissue （P<0.01）， and up-regulated protein levels of TLR4， MyD88， p-NF-κB/NF-κB， MMP-9， and VEGFA in the ectopic endometrial tissue （P<0.05， P<0.01）. Compared with the model group， all treatment groups exhibited a reduction in the ectopic lesion volume （P<0.01）. Furthermore， the writhing response scores were decreased in the high-dose Luoshi Neiyi prescription group and the dienogest group （P<0.05， P<0.01）. The pathological state of the ectopic endometrial tissue was alleviated to varying degrees in the treatment groups. Low-dose Luoshi Neiyi prescription reduced serum PGE₂ levels， and high-dose Luoshi Neiyi prescription and dienogest decreased serum IL-1β and PGE₂ levels in EMs rats （P<0.05， P<0.01）. The treatment groups showed decreased positive expression of MMP-9 and VEGFA in the ectopic endometrial tissue （P<0.05， P<0.01） and down-regulated protein levels of TLR4， MyD88， p-NF-κB/NF-κB， MMP-9， and VEGFA in the ectopic endometrial tissue （P<0.05， P<0.01）. The cell experiments showed that 5%， 10%， and 20% Luoshi Neiyi prescription-containing sera significantly reduced the viability and inhibited the migration of iheESCs. Compared with the control group， 5%， 10%， and 20% Luoshi Neiyi prescription-containing serum groups and the TAK-242 group showed reduced protein levels of TLR4， MyD88， p-NF-κB/NF-κB， MMP-9， and VEGFA （P<0.01）. Compared with the control group， the LPS group showed increased expression of the above proteins （P<0.01）. Compared with the LPS group， the LPS+5%， 10%， and 20% Luoshi Neiyi prescription-containing serum groups showed reduced expression of the above proteins （P<0.05， P<0.01）. ConclusionLuoshi Neiyi prescription may modulate the TLR4/NF-κB signaling pathway to reduce the inflammatory response and histopathological damage in the eutopic endometrium and suppress the adhesive， invasive， and angiogenic biological processes in the ectopic endometrial tissue， thereby exerting its therapeutic effect on EMs.

ObjectiveTo develop a carbon nanoparticle-manganese dioxide (CN-MnO₂) injection integrating visually identifiable staining and magnetic resonance (MR) T₁/T₂ dual-modal imaging, and to systematically evaluate its physicochemical properties, critical quality attributes, biocompatibility, and efficacy for gastrointestinal marking and localization. MethodsCN-MnO₂ was prepared using a mild liquid-phase in situ redox deposition strategy and characterized by transmission electron microscopy (TEM), dynamic light scattering (DLS), zeta potential analysis, X-ray photoelectron spectroscopy (XPS), and X-ray diffraction (XRD). The pH of CN-MnO₂ was directly measured using a pH meter. The osmolality pressure of CN-MnO₂ was determined by the freezing point depression method. The bacterial endotoxin was detected using the gel-clot Limulus amebocyte lysate test, which did not rely on absorbance readings. The appearance of the CN-MnO₂ injection solution stored at 4 ℃ under light shielding and sealed for 0, 5, and 10 days was observed, and its hydrated particle size, polydispersity index, zeta potential, and pH were also measured. With gadopentetate dimeglumine (Gd-DTPA) as the positive control, different concentration gradients were established, and T₁ and T₂ relaxation times were measured to construct concentration-dependent plots of the longitudinal and transverse relaxation rates (R₁ and R₂) and calculate the corresponding relaxivities (r₁ and r₂), compare relaxivity performance, and determine the working concentration for subsequent in vivo experiments. Cytocompatibility was assessed using the methyl thiazolyl tetrazolium (MTT) assay and live/dead staining. Sixteen male 6-week-old SD rats were randomly assigned to an experimental group and a control group (n=8 per group) and subcutaneously injected with CN-MnO₂ injection at a total Mn concentration of 0.80 mmol/L or an equal volume of normal saline, respectively. Short-term in vivo safety was evaluated by regular body weight monitoring, hematoxylin-eosin (HE) staining of major organs, hematological and serum biochemical analyses, inflammatory cytokine detection, and inductively coupled plasma optical emission spectrometry (ICP-OES) analysis. Eight male 5–6-month-old Bama minipigs were randomly assigned to a CN-MnO₂ group (n=6) and a carbon nanoparticle suspension injection (CNSI) group (n=2). CN-MnO₂ injection or CNSI was injected subserosally into the stomach, small intestine, and colon. Within 5 min after each injection, the maximum staining diameter at each injection site was measured and recorded. MR T1WI/T2WI scans were performed on days 3 and 7 after injection to evaluate MR visualization at the injection sites. At the end of observation, the injection sites and major organs were collected for histopathological evaluation by hematoxylin-eosin staining. ResultsCN-MnO₂ appeared as a black suspension, with no obvious aggregation or sedimentation after standing at room temperature (20-25 ℃) for 15 days. Its average hydrodynamic diameter was approximately (228.7±15.3) nm, the polydispersity index was 0.184±0.021, and the zeta potential was (-24.6±2.8) mV. XPS showed that CN-MnO₂ was mainly composed of C, O, and Mn, with oxygen accounting for approximately 68.82%. The Mn 2p spectrum exhibited two characteristic peaks at 642.4 and 653.1 eV. XRD indicated that low-crystallinity δ-MnO₂ was dispersed within the carbon matrix. The pH was 6.82±0.06, the osmolality was (298.4±6.7) mOsm/kg, and the bacterial endotoxin level was below the detection limit. In vitro MR imaging showed a clear concentration-dependent response, with r₁ and r₂ values of 9.6 L·mmol^-1·s^-1 and 17.2 L·mmol^-1·s^-1, respectively, indicating favorable T₁/T₂ contrast performance superior to that of Gd-DTPA. At 0.80 mmol/L, CN-MnO₂ showed no obvious cytotoxicity in vitro. Short-term safety evaluation in SD rats revealed no obvious inflammatory response, impairment of liver or renal function, or abnormal manganese accumulation in major organs. In Bama minipigs, the mean staining diameters of the CN-MnO₂ group in the stomach, small intestine, and colon were (0.65±0.12) cm, (0.61±0.11) cm, and (0.58±0.10) cm, respectively, with no significant difference among the three injection sites (P>0.05). The overall mean staining diameter was smaller in the CN-MnO₂ group than in the CNSI group (P<0.05). MR examinations showed that the injection sites were identifiable on both day 3 and day 7, with stronger signals observed on day 3. At the end of observation, no obvious pathological abnormalities were observed at the injection sites or in major organs. ConclusionA CN-MnO₂ injection with uniform particle size and stable physicochemical properties can be successfully prepared. This formulation integrates visible gastrointestinal staining with MR T₁/T₂ dual-modal imaging, providing a new strategy for preoperative gastrointestinal marking and localization as well as precise intraoperative resection.

OBJECTIVE: To summarize the early clinical results and safety of Stand-alone OLIF application of lumbar lesions, and explored its surgical indications. METHODS: Total of 92 cases of lumbar spine lesions treated with Stand-alone OLIF at two medical centers from October 2014 to December 2018 were retrospectively analyzed, including 30 males and 62 females with an average age of (61.20±12.94) years old ranged from 32 to 83 years old. There were 20 cases of lumbar spinal stenosis, 15 cases of lumbar disc degeneration, 11 cases of lumbar degenerative spondylolisthesis, 6 cases of discogenic low back pain, 7 cases of giant lumbar disc herniation, 13 cases of primary lumbar discitis, 6 cases of adjacent vertebral disease after lumbar internal fixation surgery, and 14 cases of degenerative lumbar scoliosis. Pre-operative dual energy X-ray bone density examination 31 cases' T-values ranged from -1 to -2.4, 8 cases' T-values ranged from -2.5 to -3.5, and the rest had normal bone density. The number of fusion segments: 68 cases of single segment, 9 cases of two segment, 12 cases of three segment , and 3 cases of four segment. Fusion site:L_1,2 1 case, L_2,3 4 cases, L_3,4 10 cases, L_4,5 53 cases, L_2,3-L_3,4 3 cases, L_3,4-L_4,5 6 cases, L_1,2L_2,3L_3,4 1 case, L_1,2L_3,4L_4,5 1 case, L_2,3L_3,4L_4,5 10 cases, L_1,2L_2,3L_3,4L_4,5 3 cases. The clinical results and imaging results of this group of cases were observed, as well as the complications. RESULTS: The surgical time ranged from 40 to 140 minutes with an average of (60.92±27.40) minutes. The intraoperative bleeding volume was 20 to 720 ml with an average of (68.22±141.60) ml. The patients had a follow-up period of 6 to 84 months with an average of (38.50±12.75) months. The height of the intervertebral space recovered from (9.23±1.94) mm in preoperative to (12.68±2.01) mm in postoperative, and (9.11±1.72) mm at the last follow-up, there was a statistically significant difference(F=6.641, P=0.008);there was also a statistically significant difference between the postoperative and preoperative height of the intervertebral space(t=9.27, P<0.000 1);and there was also a statistically significant difference (t=10.06, P<0.000 1) between the last follow-up and postoperative height of the intervertebral space. At the last follow-up, cage subsidence grading was as follows:level 0 in 69 cases (76 segments), levelⅠin 17 cases (43 segments), level Ⅱin 5 cases (14 segments), and level Ⅲ in 1 case (1 segment);according to the number of segments, normal subsidence accounts for 56.72%, abnormal subsidence accounts for 43.28%. Bone mineral desity of normal subsidence groups was -0.50±0.07 whinch was better than that the abnormal subsidence groups -2.10±0.43, and the difference was statistically significant(χ²=2.275, P=0.014). As well as there was a statistically significant difference in the patient's VAS of backache from (6.28±2.11) in preoperative to (1.48±0.59) in last follow-up(t=8.56, P<0.05). The ODI recovered from (36.30±7.52)% before surgery to (10.20±2.50)% at the last follow-up, with a statistically significant difference (t=7.79, P<0.000 1). Complications involved 4 cases of intraoperative vascular injury, 21 cases of endplate injury, and 4 cases of combined vertebral fractures. The incision skin has no necrosis or infection. There were 4 cases of left sympathetic chain injury, 4 cases of transient left hip flexion weakness, 2 cases of left thigh anterolateral numbness with quadriceps femoris weakness, and 1 case of incomplete intestinal obstruction;8 cases were treated with posterior pedicle screw fixation due to fusion cage settlement accompanied by stubborn lower back pain, and 6 cases were treated with fusion cage settlement and lateral displacement. According to the actual number of cases, there were 38 complications, with an incidence rate of 41.3%. CONCLUSION The application of Stand alone OLIF in lumbar spine disease fusion has achieved good early results, with obvious clinical advantages, but also there are high probability of complications. It is recommended to choose carefully. It is necessary to continuously summarize and gradually clarify and complete the surgical indications and specific case selection criteria.
Humans ; Male ; Female ; Middle Aged ; Spinal Fusion/methods* ; Lumbar Vertebrae/injuries* ; Aged ; Adult ; Retrospective Studies ; Aged, 80 and over

Several types of arthritis share the common feature that the generation of inflammatory mediators leads to joint cartilage degradation. However, the shared mechanism is largely unknown. H2BK120ub1 was reportedly involved in various inflammatory diseases but its role in the shared mechanism in inflammatory joint conditions remains elusive. The present study demonstrated that levels of cartilage degradation, H2BK120ub1, and its regulator WW domain-containing adapter protein with coiled-coil (WAC) were increased in cartilage in human rheumatoid arthritis (RA) and osteoarthritis (OA) patients as well as in experimental RA and OA mice. By regulating H2BK120ub1 and H3K27me3, WAC regulated the secretion of inflammatory and cartilage-degrading factors. WAC influenced the level of H3K27me3 by regulating nuclear entry of the H3K27 demethylase KDM6B, and acted as a key factor of the crosstalk between H2BK120ub1 and H3K27me3. The cartilage-specific knockout of WAC demonstrated the ability to alleviate cartilage degradation in collagen-induced arthritis (CIA) and collagenase-induced osteoarthritis (CIOA) mice. Through molecular docking and dynamic simulation, doxercalciferol was found to inhibit WAC and the development of cartilage degradation in the CIA and CIOA models. Our study demonstrated that WAC is a key factor of cartilage degradation in arthritis, and targeting WAC by doxercalciferol could be a viable therapeutic strategy for treating cartilage destruction in several types of arthritis.

Traditional development of small protein scaffolds has relied on display technologies and mutation-based engineering, which limit sequence and functional diversity, thereby constraining their therapeutic and application potential. Protein design tools have significantly advanced the creation of novel protein sequences, structures, and functions. However, further improvements in design strategies are still needed to more efficiently optimize the functional performance of protein-based drugs and enhance their druggability. Here, we extended an evolution-based design protocol to create a novel minibinder, BindHer, against the human epidermal growth factor receptor 2 (HER2). It not only exhibits super stability and binding selectivity but also demonstrates remarkable properties in tissue specificity. Radiolabeling experiments with ^99mTc, ⁶⁸Ga, and ¹⁸F revealed that BindHer efficiently targets tumors in HER2-positive breast cancer mouse models, with minimal nonspecific liver absorption, outperforming scaffolds designed through traditional engineering. These findings highlight a new rational approach to automated protein design, offering significant potential for large-scale applications in therapeutic mini-protein development.

Aim To investigate the effect of linarin on improving cognitive behavior of APP/PS1 mice,and to explore the therapeutic effect of linarin on A β deposi-tion and neuroinflammation and its correlation.Meth-ods APP/PS1 transgenic mice were randomly divid-ed into the model group,high-dose group,medium-dose group,low-dose group and positive control group.C57BL/6J mice were set as the normal group.Morris water maze was used to evaluate the learning and mem-ory abilities of mice.TUNEL staining was used to de-tect the apoptosis of neurons in the CA1 region of mice.IHC was used to detect the expression levels of Aβ42 and GFAP.Western blot was used to detect the expression levels of BACE1 and PS-1.Results Com-pared with the normal group,mice of the model group showed lower NCP,shorter target quadrant travel,less target quadrant residence time percentage(all P＜0.01),higher apoptosis rate of neurons in the CA1 re-gion(P＜0.01),significantly higher protein expres-sion levels of A β42 and GFAP(all P＜0.01),and significantly higher protein expression levels of BACE1 and PS-1(all P＜0.01).Compared with the model group,the medium-dose group,high-dose group and positive control group showed higher NCP,longer tar-get quadrant travel,more target quadrant residence time percentage(all P＜0.05),lower apoptosis rate of neurons in the CA1 region(P＜0.01),significantly lower protein expression levels of A β42 and GFAP(all P＜0.01),and significantly lower protein expression levels of BACE1 and PS-1(all P＜0.01).Conclu-sions Linarin can inhibit two key enzymes to reduce the decomposition of APP and the generation of A β42,thereby inhibiting the activation of astrocytes,allevia-ting neuroinflammation,improving the core pathologi-cal features of AD,and thus significantly improving learning and memory impairment in APP/PS1 mice.