Lysosome, the digestive organelle in eukaryotic cells, plays an important role in the degradation and recirculation of cellular products as well as in maintaining the stability of cellular metabolic microenvironment. Surface-enhanced Raman scattering (SERS) is a molecular fingerprint technology with high detection sensitivity and photostability, suited for revealing various intracellular molecular information by inducing endocytosis of SERS-active nanoparticles. However, it remains challenging to selectively extract the molecular information of specific organelles (e.g., lysosomes) from a high-dimensional spectral set. Herein, we proposed a novel paradigm by combining label-free SERS spectroscopy with confocal fluorescence imaging to investigate the digestion behavior of lysosomes in cells. The structural similarity algorithm was innovatively introduced and exhibited its effectiveness in screening out the wavenumbers in the SERS spectral set with high correlation with the metabolic behaviors of lysosomes. With comprehensive experiments on HeLa single cells, we captured the intracellularmacromolecular digestion phenomenon and discovered the changing pattern of cellular SERS spectra after starvation-induced autophagy, and analyzed the molecular information within the lysosomes in three-dimensional space.

Liver fibrosis is a major risk factor for hepatocellular carcinoma origin, and its progression not only correlates with oxidative stress and inflammation, but also is encouraged by autophagy hold-up. Therefore, new solutions to effectively attenuate oxidative stress and inflammation and coincidently favor autophagy are highly demanded to reverse liver fibrosis, and even hamper its escalation into hepatocellular carcinoma. Herein, the porous manganese-substituted Prussian blue (PMPB) analogs are harnessed to activate autophagy, scavenge reactive oxygen species (ROS), and suppress inflammation for liver fibrosis therapy. PMPB can effectively inhibit macrophage activation, facilitate macrophage autophagy, eradicate ROS, and blockade cellular cross-talk, thus impeding further inflammation progression. Moreover, the favorable spontaneous capture of PMPB by Kupffer cells allows more PMPB accumulation in liver to significantly attenuate liver injury and collagen deposition, thereby inhibiting the progression of liver fibrosis. PMPB-based nanomedicine shows great potentials in promoting autophagy activation, eliminating ROS, inhibiting inflammation, and protecting hepatocytes from oxidative stress-arised damages, which eventually attenuate the extent of liver fibrosis, holding great promise in clinical translation for treating liver fibrosis.

Despite progression in advanced treatments for malignant tumors, surgery remains the primary treatment intervention, which removes a large portion of firm tumor tissues; however, the postoperative phase poses a possible risk for provincial tumor recurrence and metastasis. Consequently, the prevention of tumor recurrence and metastasis has attracted research attention. In this review, we summarized the postoperative treatment strategies for various tumors from both basic research and clinical perspectives. We delineated the underlying factors contributing to the recurrence of malignant tumors with a substantial prevalence rate, related molecular mechanisms of tumor recurrence post-surgery, and relatedmeans of monitoring recurrence andmetastasis after surgery. Furthermore, we described relevant therapeutic approaches for postoperative tumor recurrence, including chemotherapy, radiation therapy, immunotherapy, targeted therapy, and photodynamic therapy. This review focused on the emerging technologies used for postoperative tumor treatment in recent years in terms of functional classification, including the prevention of postoperative tumor recurrence, functional reconstruction, and monitoring of recurrence. Finally, we discussed the future development and deficiencies of postoperative tumor therapy. To understand postoperative treatment strategies for tumors from clinical treatment and basic research and further guide the research directions for postoperative tumors.

Nano-sized polymer systems are often used as carriers for drugs and contrast agents to increase circulation time and solubility and to reduce possible side effects. These nanomedicines usually accumulate in tumor tissue due to the enhanced permeability and retention (EPR) effect. However, a targeting group may be attached to the polymer carrier in addition to the active substance to further increase tumor accumulation and specificity. In this study, the oligopeptide sequence RGD was chosen to target α_vβ₃ integrins overexpressed in the tumor vasculature and on some tumor cells. A set of polymer conjugates bearing a fluorescent dye and RGD peptide of different structures (linear, cyclic, branched) was prepared for use in tumor diagnosis, with a potential future application in navigated surgery. The accumulation of the most promising candidate, a targeted fluorescent nanoprobe, increased by 35%in glioblastoma tumors compared to the non-targeted control, which accumulated only due to the EPR effect. However, the administration of a polymer-boundmodified cilengitide as an antiangiogenic treatment did not show a beneficial effect in the suppression of angiogenesis.

Mycobacterium tuberculosis (MTB) is the causal pathogen of tuberculosis (TB). Rapid and accurate detection of live MTB is important for transmission control and patient treatment. Here, we described a clustered regularly interspaced short palindromic repeat (CRISPR)-Cas13a-based molecular diagnosis approach for rapid and specific detection of live MTB. This detection method, which we termed CRISPR-Live-MTB, contained two consecutive reactions including nuclear acid sequence-based amplification (NASBA) and CRISPR-Cas13a collateral cleavage reaction. CRISPR-Live-MTB could efficiently detect MTB single-stranded RNA (ssRNA) in 2 hours with high specificity over double-stranded DNA (dsDNA). Importantly, CRISPR-Live-MTB exhibited a limit of detection of 2.4 copies for MTB ssRNA, which was 1000 times lower than that of the clinically used NASBA method. Moreover, lateral flow was integrated into the CRISPR-Live-MTBmethod to enable point-of-care testing application with a sensitivity of 95% and a specificity of 100%. Overall, our study demonstrated the feasibility of CRISPR-Live-MTB as a rapid, sensitive, and specific approach for live MTB detection.

Fluorescent dyes that emit in the second near-infrared (NIR-II, 1000–3000 nm) region have provided significant advances toward real-time and high-resolution imaging of vessel and lymphatic system. However, in vivo NIR-II tracking of the fate of labeled cells still remains challenging. Here, we develop a shielding unit–donor–acceptor–donor–shielding unit (S-D-A-D-S) NIR-II fluorophore (FE-4ZW) with zwitterionic terminal groups for high-efficiency cell labeling without using cell-penetrating peptides, which provides for enhanced noninvasive in vivo determination of the location of cell migration. The tethering terminal sulfoammonium inner salts are featured with its high affinity for cell membranes, thereby enabling the stable labeling even for fixed cells. The fate of transplanted stem cell and the tumor cell migration along lymphatic system in brain or periphery tissues are clearly monitored by the cell-internalized FE-4ZW. We also confirmed that a clinically used surfactant, D-α-tocopheryl polyethylene glycol-1000 succinate, can reduce the liver and spleen uptake of FE-4ZW. The fluorophore design strategy and cell-labeling technology reported here open a new realm in the visualization of cell migration and insight into the relocation process, thereby ultimately providing an opportunity to investigate in greater detail of the underlying mechanisms of stem cell therapy and tumor metastasis.

Testicular microcirculation is closely related to spermatogenic function and seminiferous tubular function. The diagnosis and monitoring of testicular diseases can be associated with testicular microcirculation; however, there are currently no effective non-invasive methods for super-resolution imaging of testicular microcirculation. In this study, we introduced state-of-the-art graph-based tracking with the Kalman motion model algorithm to non-invasively image human testicular microcirculation for the first time with a regular frame-rate clinical ultrasound imaging system (37 Hz). Two distinct testicular vessels with an 81 µm separation were resolved in the testicular vasculature, surpassing all other imaging modalities. In a retrospective study, we performed contrast-enhanced ultrasound examinations(CEUS) and ultrasound localizationmicroscopy (ULM) processing on the included 76 infertile patients and 15 healthy controls from August 2021 to May 2023 and obtained super-resolution images of testicular microcirculation with sub-diffraction resolution. Through the results of one-way analysis of variance tests and receiver operating characteristic analyses,we found that the ULM-based parameters hold promise as clinical guidance for differentiating between non-obstructive and obstructive male infertility. The mean vessel diameter achieved an area under the curve (AUC) of 0.920 (95%confidence interval [CI]: 0.847–0.994, p < .001) with a cut-off value of 170.9 µm in oligoasthenospermia, and an AUC of 0.952 (95% CI: 0.875–1.000, p < .001) with a cut-off value of 169.9 µm in azoospermia patients, respectively, addressing a significant clinical challenge.

Transcription factor 21 (TCF21) and estrogen receptor beta (ERβ, encoded by ESR2) are highly expressed in endometriotic stromal cells (ESCs) and contribute to the pathogenesis of endometriosis. However, the exploration of TCF21 and ERβ expression regulation at the molecular level remains limited. Here, by using bioinformatics analysis and experimental verification, we identified PES1, also known as Pescadillo, as a negative regulator in the development of endometriosis that downregulates TCF21 and ERβ expression in ESCs. PES1 overexpression regulated critical biological processes involved in endometriosis development, such as invasion and apoptosis. A coimmunoprecipitation assay showed that PES1 could form a complex with Forkhead box M1 (FOXM1). Further analyses elucidated that siPES1 in ectopic lesions decreased the stability of FOXM1 protein and reduced the binding activities of FOXM1 to TCF21 and ESR2 promoters, thus weakening the transcriptional inhibition of TCF21 and ERβ by FOXM1. Moreover, in an endometriosis mousemodel, overexpressing PES1 effectively reduced the growth of ectopic lesions and suppressed TCF21 and ERβ expression, which suggests a promising therapeutic strategy for endometriosis. Collectively, our results indicate that the loss of PES1 in ectopic lesions contributes to endometriosis progression by upregulating ERβ and TCF21 expression through heterodimer formation with FOXM1. Moreover, targeting PES1 could serve as a treatment method for endometriosis.

Tumor-derived exosomes are crucial for early non-invasive and accurate tumor diagnosis in clinical diagnostics. The development of highly sensitive, simple, and intuitive exosome assays has sparked a research upsurge in clinical diagnostics. Here, we develop a bio-responsive intelligent DNA hydrogel loaded with CRISPR/Cas12a for universal and ultrasensitive detection of the exosomes. The aptamer serves as the target response unit and switch, competitively disintegrating the region of the DNA linkers and then Cas12a/crRNA was released and activated, resulting in a high fluorescent intensity for exosome detection at the detection limit of 119 particles/µL. Moreover, a constructed colorimetric tube is made by loading a colorimetric filter membrane on the tube lid and intelligent DNA hydrogel on the tube bottom, which enables one-pot portable colorimetric detection. Without the need for laboratory instruments and professionals, this strategy allows for naked eye detection with limit of detection as low as 10⁴ particles/µL, and shows great applicability in distinguishing between healthy individuals, pretreatment patients, and post-treatment patients after obtaining a testable analyte. In this study, an ultrasensitive detection platform for exosomes that enables one-step sensing and dual signal output was introduced. The findings here suggest that this platform is a promising tool for the application of liquid biopsy based on exosomes in clinical diagnosis.

Surface-enhanced Raman spectroscopy (SERS) has become an essential biodetection technique. Due to its high sensitivity, good signal specificity, and resistance to photobleaching, SERS has been widely used in biomedical research fields such as molecular imaging, tumor diagnosis, and drug monitoring. This review focuses on the progress of SERS in biomedical applications.We first introduce the basic principle of SERS and the progress of substrate research. Then, we summarize the latest research progress on SERS in drugmonitoring, cell and exosome detection, tumor imaging, and detection platforms combining microfluidic and lateral flow technologies. Subsequently, the applied research of SERS in early diagnosis of pancreatic cancer and drug efficacy monitoring is described. Finally, the future development direction and possible challenges of SERS in tumor diagnosis and treatment are proposed.

Cartilage defects resulting from injury or degeneration are a common clinical problem, and due to its avascular nature, articular cartilage has poor self-healing capacity. Three-dimensional (3D) bioprinting has attracted great attention in tissue engineering. Melatonin (MT), a hormone mainly secreted at night, plays an important role in tissue repair. Small extracellular vesicles (sEV) are considered ideal drug delivery vehicles and MT-sEV (sleep-related sEV) have the potential ability to promote cartilage regeneration. Here, biomimetic multilayer scaffolds were fabricated using 3D bioprinting. A double network hydrogel, composed of methacrylated hyaluronic acid and gelatin methacryloyl (HG), was prepared. MT-sEV and HG hydrogel were used to create a cartilage layer. A bone layer was formed using poly(ϵ-caprolactone) and hydroxyapatite ultralong nanowires. Additionally, two bioinks were alternately printed at the interface layer. The results of RNA sequencing revealed the potential regulatory mechanisms. MTsEV showed promotional effects on cell migration, proliferation, chondrogenic differentiation, and extracellular matrix (ECM) deposition. Moreover, MT-sEV altered macrophage polarization and regulated the expression of inflammatory cytokines. In vivo experiments demonstrated that the biomimetic multilayer scaffolds promoted cartilage regeneration. These experiments demonstrated the ability of MT-sEV to regulate the immune microenvironment and promote the secretion of ECM, providing a promising strategy for cartilage regeneration.

Atomic force microscopy (AFM) is an analytical technique that is increasingly utilized to determine interaction forces on the colloidal and cellular level. Fluidic force microscopy, also called FluidFM, became a vital tool for biomedical applications. FluidFM combines AFM and nanofluidics by means of a microchanneled cantilever that bears an aperture instead of a tip at its end. Thereby, single colloids or cells can be aspirated and immobilized to the cantilever, for example, to determine adhesion forces. To allow for quantitative measurements, the socalled (inverse) optical lever sensitivity (OLS and InvOLS, respectively) must be determined, which is typically done in a separate set of measurements on a hard, non-deformable substrate. Here, we present a different approach that is entirely based on hydrodynamic principles and does make use of the internal microfluidic channel of a FluidFM-cantilever and an external pressure control. Thereby, a contact-free calibration of the (inverse) optical lever sensitivity (InvOLS) becomes possible in under a minute. A quantitative model based on the thrust equation, which is well-known in avionics, and finite element simulations, is provided to describe the deflection of the cantilever as a function of the externally applied pressure. A comparison between the classical and the here-presented hydrodynamic method demonstrates equal accuracy.

DNA programming, which is based on the principle of base complementary pairing and Boolean operations, exhibits organizational structures and algorithms similar to those observed in machine language. Consequently, the practical implementation of DNA logic programming can be achieved through the utilization of programming techniques, enabling the discrimination and output generation. In recent years, DNA programming has witnessed a convergence with disciplines, such as life sciences, medicine, and other interdisciplinary areas, thereby giving rise to an advanced research system that yields valuable insights. This development has paved the way for multidisciplinary cutting-edge research. Furthermore, the successful transition from conceptualization to the practical implementation of DNA programming has been accomplished. This review summarizes the recent advances in DNA logic programming within the biomedical fields, specifically emphasizing the conceptualization and execution of DNA logic programming constructs. The benefits and obstacles associated with the adoption of DNA programming in cutting-edge research areas are also highlighted.

It is widely recognized that platinum-based chemotherapy, particularly cisplatin therapy, can cause ototoxicity. At present, there are no Food and Drug Administration-approved drugs to prevent or alleviate ototoxicity. Ototoxicity is generally believed to be caused by excessive reactive oxygen species production in the inner ear. Accordingly, a variety of antioxidants have been developed to protect against ototoxicity. To improve the efficiency of drug delivery to the cochlea, here, we synthesized simple and easy-to-obtain glutathione carbon dots (GSH CDs) with ultra-small dimensions. The experimental results revealed that the GSH CDs have strong free-radical scavenging activity and can restore mitochondrial function, maintain hair cell stability, and protect hair cells from cisplatin-induced oxidative stress. Thus, GSH CDs may serve as a new therapeutic agent for preventing cisplatin-induced ototoxicity.

Malfunction of neutrophil apoptosis and elevated serum lactate levels are the key cellular mechanism of immune suppressive status in septic patients. However, whether increased lactate affects apoptosis of neutrophils and aggravates sepsis development, and the molecular mechanism remain unknown. In this study, first, we analyzed the transcriptional profiles of blood cells in sepsis patients(n = 39) and healthy volunteers (n = 40) to reveal that there is close correlation between the lactate-related gene expression changes associated with lactate production and immune function in leukocytes, especially in neutrophils. Further, we explored the close relationship between lactate and delayed neutrophil apoptosis in human neutrophils. Programmed cell death 1 leg and (PD-L1) was highly expressed in septic patients compared with healthy volunteers. Then, we indicated that elevated levels of lactate in human neutrophils decreased neutrophil apoptosis (P < .001) by up regulating PD-L1 expression. Inhibition of monocarboxylate transporter 1 (MCT1) significantly attenuated neutrophil apoptosis caused by lactate (P < .001). We further performed in vivo experiments in sepsis mice model and determined that increased lactate decreased neutrophil apoptosis (P < .05) and reduces mice survival rate (P < .001), which could also be rescued by MCT1 inhibitor (P < .05). This study revealed that elevated level of lactate in sepsis upregulates PD-L1 expression to decrease apoptosis throughMCT1 in neutrophils, which provides new insight into sepsis treatment strategy by reducing lactate accumulation.