siRNA-loaded folic acid-modified TPGS alleviate MASH via targeting ER stress sensor XBP1 and reprogramming macrophages.

Macrophages show high plasticity and play a vital role in the progression of metabolic dysfunction-associated steatohepatitis (MASH). X-box binding protein 1 (XBP1), a key sensor of the unfolded protein response, can modulate macrophage-mediated pro-inflammatory responses in the pathogenesis of MASH. However, how XBP1 influences macrophage plasticity and promotes MASH progression remains unclear. Herein, we formulated an Xbp1 siRNA delivery system based on folic acid modified D-α-tocopheryl polyethylene glycol 1000 succinate nanoparticles (FT@XBP1) to explore the precise role of macrophage-specific Xbp1 deficiency in the progression of MASH. FT@XBP1 was specifically internalized into hepatic macrophages and subsequently inhibited the expression of spliced XBP1 both in vitro and in vivo. It promoted M1-phenotype macrophage repolarization to M2 macrophages, reduced the release of pro-inflammatory factors, and alleviated hepatic steatosis, liver injury, and fibrosis in mice with fat-, fructose- and cholesterol-rich diet-induced MASH. Mechanistically, FT@XBP1 promoted macrophage polarization toward the M2 phenotype and enhanced the release of exosomes that could inhibit the activation of hepatic stellate cells. A promising macrophage-targeted siRNA delivery system was revealed to pave a promising strategy in the treatment of MASH.

Programmable Framework Nucleic Acid-Modified Nanomagnetic Beads for Efficient Isolation of Exosomes and Exosomal Proteomics Analysis.

Exosomes are increasingly being regarded as emerging and promising biomarkers for cancer screening, diagnosis, and therapy. The downstream molecular analyses of exosomes were greatly affected by the isolation efficiency from biosamples. Among the current exosome isolation strategies, affinity nanomaterials performed comparably better with selectivity and specificity. However, these techniques did not take the structure and size of exosomes into account, which may lead to a loss of isolation efficiency. In this article, a framework nucleic acid was employed to prepare a well-designed nanosized bead Fe3O4@pGMA@DNA TET@Ti4+ for enrichment of exosomes. The abundant phosphate groups in the framework nucleic acid provide binding sites to immobilize Ti4+, and its rigid three-dimensional skeleton makes them act as roadblocks to barricade exosomes and provide affinity interactions on a three-dimensional scale, resulting in the improvement of isolation efficiency. The model exosomes can be effectively isolated with 92% recovery in 5 min. From 100 µL of HeLa cell culture supernatant, 34 proteins out of the top 100 commonly identified exosomal proteins were identified from the isolated exosomes by the novel beads, which is obviously more than that by TiO2 (19 proteins), indicating higher isolation efficiency and exosome purity by Fe3O4@pGMA@DNA TET@Ti4+ beads. The nanobeads were finally applied for comparing exosomal proteomics analysis from real clinical serum samples. Twenty-five upregulated and 10 downregulated proteins were identified in the lung cancer patients group compared to the health donors group, indicating that the novel nanobeads have great potential in isolation of exosomes for exosomal proteomics analysis in cancer screening and diagnosis.

Case report: A case report of co-morbidity of cervical intraepithelial neoplasia III and urethral cancer associated with HPV16.

In this report, we present a case of a woman with concurrent cervical intraepithelial neoplasia grade III (CIN III) and urethral cancer, both associated with HPV16 infection. This unique case was initially brought to attention due to postmenopausal vaginal bleeding, despite the absence of urological symptoms and negative tumor markers. An unexpected discovery of pelvic lymph node metastasis during a hysterectomy intended for CIN III highlighted the rare coexistence of these conditions, with urethral cancer also linked to HPV-16 within the urethral lesion. This case emphasizes the diagnostic challenges faced by HPV-related cervical lesions and the critical need for increased vigilance, even when urological symptoms are not apparent. The findings underline the potential complexity of HPV-associated lesions and advocate for comprehensive screening strategies to ensure the timely detection and management of such intricate cases.

Multifunctional Bispecific Nanovesicles Targeting SLAMF7 Trigger Potent Antitumor Immunity.

The effectiveness of immune checkpoint inhibitor (ICI) therapy is hindered by the ineffective infiltration and functioning of cytotoxic T cells and the immunosuppressive tumor microenvironment (TME). Signaling lymphocytic activation molecule family member 7 (SLAMF7) is a pivotal co-stimulatory receptor thought to simultaneously trigger NK-cell, T-cell, and macrophage antitumor cytotoxicity. However, the potential of this collaborative immune stimulation in antitumor immunity for solid tumors is underexplored due to the exclusive expression of SLAMF7 by hematopoietic cells. Here, we report the development and characterization of multifunctional bispecific nanovesicles (NVs) targeting SLAMF7 and glypican-3-a hepatocellular carcinoma (HCC)-specific tumor antigen. We found that by effectively "decorating" the surfaces of solid tumors with SLAMF7, these NVs directly induced potent and specific antitumor immunity and remodeled the immunosuppressive TME, sensitizing the tumors to programmed cell death protein 1 (PD1) blockade. Our findings highlight the potential of SLAMF7-targeted multifunctional bispecific NVs as an anticancer strategy with implications for designing next-generation targeted cancer therapies.

Development of cancer biomarker heat shock protein 90α certified reference material using two different isotope dilution mass spectrometry techniques.

Heat shock protein 90α (HSP90α) has been regarded as an important indicator for judging tumor metastasis and prognosis due to its significant upregulation in various tumors. Therefore, the accurate quantification of HSP90α is of great significance for clinical diagnosis and therapy of cancers. However, the lack of HSP90α certified reference material (CRM) leads to the accuracy and consistency of quantification methods not being effectively evaluated. Besides, quantitative results without traceability make comparisons between different studies difficult. In this study, an HSP90α solution CRM was developed from the recombinant protein raw material. The recombinant protein is a dimer, and the purity of the CRM candidate reached 96.71%. Both amino acid analysis-isotope dilution mass spectrometry (AAA-IDMS) and unique peptide analysis-isotope dilution mass spectrometry (UPA-IDMS) were performed to measure the content of HSP90α in the solution CRM candidate, and the certified value was assessed to be 66.2 ± 8.8 µg/g. Good homogeneity of the CRM was identified, and the stability examination suggested that the CRM was stable for at least 4 months at - 80 °C and for 7 days at 4 °C. With traceability to SI unit (kg), this CRM has potential to help establish a metrological traceability chain for quantification of HSP90α, which will make the quantification results standardized and comparable regardless of the quantitative methods.

Natural deep eutectic solvent-A novel green solvent for protein stabilization.

Natural deep eutectic solvents (NADESs) have been explored to provide a favorable environment for protein stabilization. In this context, NADESs were prepared with the molar ratio of trehalose to betaine ranging from 1:3 to 1:9 (NADES 1-3 to NADES 1-9). There was a strong hydrogen bond interaction between trehalose and betaine, and the interaction weakened with the reduction of trehalose. The NADES 1-7 had good thermal stability (-60-100 °C), low viscosity, and suitable pH (around 7). Trypsin had the highest relative enzyme activity in 50 % (v/v) NADES 1-7 under different temperatures, pH, and storage time. Furthermore, the changes in kinetic parameters indicated that the hydrogen bond environment of 50 % NADES 1-7 increased the contact between the substrate and the trypsin, speeding up the enzymatic reaction rate. This stabilizing effect mainly derived from the virtue of NADES 1-7 itself rather than the superposition of individual components. Additionally, spectral analysis revealed that the NADES 1-7 promoted trypsin conformational folding, effectively protecting the natural structure of trypsin. Importantly, the NADES 1-7 had good biocompatibility, further expanding its application.

Corneal Epithelial Remodeling in a 6-Month Follow-up Period in Myopic Corneal Refractive Surgeries.

PURPOSE: To investigate corneal epithelial thickness changes during a 6-month follow-up period after transepithelial photorefractive keratectomy (tPRK), femtosecond laser-assisted laser in situ keratomileusis (FS-LASIK), and small incision lenticule extraction (SMILE). METHODS: This prospective study included 76 eyes of 76 participants who underwent myopic refractive surgery (23 FS-LASIK, 22 SMILE, and 31 tPRK). Epithelial thickness and anterior curvature were averaged over 4 regions (subdivided into 25 areas) and measured by spectral-domain optical coherence tomography and Scheimpflug tomography before the operation (pre) and at 1 or 3 days (pos1-3d), 1 week (pos1w), and 1 month (pos1m), 3 months (pos3m), and 6 months (pos6m) postoperatively. RESULTS: The epithelial thickness of the three groups was similar in both the pre and pos6m (all P > .05), but the tPRK group fluctuated the most during the follow-up period. The largest increase was in the inferior-temporal paracentral area (7.25 ± 2.58 µm for FS-LASIK; 5.79 ± 2.41 µm for SMILE; 4.88 ± 5.84 µm for tPRK; all P < .001). Only the epithelial thickness of tPRK increased from pos3m to pos6m (P < .05), whereas all changes for FS-LASIK and SMILE were not significant (P > .05). A positive correlation of thickness changes with curvature gradient in the paracentral region of tPRK was found (r = 0.549, P = .018), but not in other regions in all groups. CONCLUSIONS: Epithelial remodeling followed different trends after different surgeries from the early postoperative stage onward, but exhibited similar values at pos6m. Although remodeling after FS-LASIK and SMILE stabilized by pos3m, it remained unstable at pos6m after tPRK. These changes may affect corneal profile and lead to deviation from the intended surgical outcome. [J Refract Surg. 2023;39(3):187-196.].

Biomechanical Effects of tPRK, FS-LASIK, and SMILE on the Cornea.

Purpose: The objective of this study is to evaluate the in vivo corneal biomechanical response to three laser refractive surgeries. Methods: Two hundred and twenty-seven patients who submitted to transepithelial photorefractive keratectomy (tPRK), femtosecond laser-assisted in-situ keratomileusis (FS-LASIK), or small-incision lenticule extraction (SMILE) were included in this study. All cases were examined with the Corvis ST preoperatively (up to 3 months) and postoperatively at 1, 3, and 6 months, and the differences in the main device parameters were assessed. The three groups were matched in age, gender ratio, corneal thickness, refractive error corrections, optical zone diameter, and intraocular pressure. They were also matched in the preoperative biomechanical metrics provided by the Corvis ST including stiffness parameter at first applanation (SP-A1), integrated inverse radius (IIR), deformation amplitude (DA), and deformation amplitude 2 mm away from apex and the apical deformation (DARatio2mm). Results: The results demonstrated a significant decrease post-operation in SP-A1 and significant increases in IIR, DA, and DARatio2mm (p < 0.05), all of which indicated reductions in overall corneal stiffness. Inter-procedure comparisons provided evidence that the smallest overall stiffness reduction was in the tPRK group, followed by the SMILE, and then the FS-LASIK group (p < 0.05). These results remained valid after correction for the change in CCT between pre and 6 months post-operation and for the percentage tissue altered. In all three surgery groups, higher degrees of refractive correction resulted in larger overall stiffness losses based on most of the biomechanical metrics. Conclusion: The corneal biomechanical response to the three surgery procedures varied significantly. With similar corneal thickness loss, the reductions in overall corneal stiffness were the highest in FS-LASIK and the lowest in tPRK.

Changes in Corneal Biomechanical Properties in PRK Followed by Two Accelerated CXL Energy Doses in Rabbit Eyes.

PURPOSE: To evaluate whether photorefractive keratectomy (PRK) combined with the two commonly delivered energy doses in accelerated corneal cross-linking (A-CXL) could help the cornea maintain its preoperative stiffness level. METHODS: A total of 72 corneas of 36 healthy white Japanese rabbits were randomly divided into four equal groups. The groups included an untreated control group and three that had undergone PRK. After tissue ablation, one of the latter three groups (PRK group) was left untreated, whereas the other two were exposed to riboflavin (0.22% concentration by volume) and ultraviolet-A (370 nm) with the same irradiation (30 mW/cm2) but different CXL energy doses of 1.8 J/cm2 (PXL group) and 2.7 J/cm2 (PXH group). Dynamic Scheimpflug analyzer (Corvis ST; Oculus Optikgeräte GmbH) measurements of stiffness parameter at first applanation (SP-A1), Stress-Strain Index (SSI), and other dynamic corneal response parameters were taken 3 days preoperatively and 1 month postoperatively. Subsequently, ex vivo inflation testing was performed and the tangent modulus of each specimen was estimated using an inverse analysis process. RESULTS: In comparison to the control group, the tangent modulus at a stress of 10 kPa decreased by 8.9% in the PRK group and increased by 10.6% and 22.4% in the PXL and PXH groups, respectively. SP-A1 decreased postoperatively in the PRK group (P < .05), indicating an overall stiffness reduction of -7.4, -3.5, and -5.3 mm Hg/mm in PRK, PXL, and PXH groups, respectively. The material stiffness parameter SSI remained almost unchanged in the PRK group (P = .989), increased slightly in the PXL group (8.3%, P = .077), and increased significantly in the PXH group (11.1%) (P < .05). CONCLUSIONS: Biomechanical deterioration following PRK was significant and could not be fully compensated for by ACXL with either 1.8 or 2.7 J/cm2 doses. The increased value of corneal overall stiffness was higher in A-CXL with 2.7 J/cm2 energy than with 1.8 J/cm2 energy. [J Refract Surg. 2021;37(12):853-860.].

Learning non-local perfusion textures for high-quality computed tomography perfusion imaging.

Background. Computed tomography perfusion (CTP) imaging plays a critical role in the acute stroke syndrome assessment due to its widespread availability, speed of image acquisition, and relatively low cost. However, due to its repeated scanning protocol, CTP imaging involves a substantial radiation dose, which might increase potential cancer risks.Methods. In this work, we present a novel deep learning model called non-local perfusion texture learning network (NPTN) for high-quality CTP imaging at low-dose cases. Specifically, considering abundant similarities in the CTP images, i.e. latent self-similarities within the non-local region in the CTP images, we firstly search the most similar pixels from the adjacent frames within a fixed search window to obtain the non-local similarities and to construct non-local textures vector. Then, both the low-dose frame and these non-local textures from adjacent frames are fed into a convolution neural network to predict high-quality CTP images, which can help better characterize the structure details and contrast variants in the targeted CTP image rather than simply utilizing the targeted frame itself. The residual learning strategy and batch normalization are utilized to boost the performance of the convolution neural network. In the experiment, the CTP images of 31 patients with suspected stroke disease are collected to demonstrate the performance of the presented NPTN method.Results. The results show the presented NPTN method obtains superior performance compared with the competing methods. From numerical value, at all dose levels, the presented NPTN method has achieved around 3.0 dB improvement of average PSNR, an increase of around 1.4% of average SSIM, and a decrease of around 4.8% of average RMSE in the low-dose CTP reconstruction task, and also has achieved an increase of around 3.4% of average SSIM and a decrease of around 61.1% of average RMSE in the cerebral blood flow (CBF) estimation task.Conclusions. The presented NPTN method can obtain high-quality CTP images and estimate high-accuracy CBF map by characterizing more structure details and contrast variants in the CTP image and outperform the competing methods at low-dose cases.

[A new cytotoxic isoflavane from Dalbergiae Odoriferae Lignum].

Fourteen compounds were isolated from the ethanol extract of Dalbergiae Odoriferae Lignum by various chromatographic techniques, including column chromatographies on silica gel, Sephadex LH-20 and semi-preparative HPLC. Their structures were determined by spectroscopic techniques as S-3'-hydroxy-7,2',4'-trimethoxyisoxane(1), 2-(2',4'-dimethoxyphenyl)-6-hydroxybenzofuran(2), 2-(2'-hydroxy-4'-methoxyphenyl)-6-methoxybenzofuran(3), 7,2',4'-trimethoxydihydroisoflavone(4), sativanone(5), 3,9-dimethoxy-6H-benzofuro[3,2-c]chromen-6-one(6),(6 aS,11 aS)-homopterocarpin(7),(6 aS,11 aS)-8-hydroxy-3,9-dimethoxypterocarpan(8),(6 aS,11 aS)-3,8,9-trimethoxypterocarpan(9), isodalbergin(10), isoliquiritigenin(11), butein(12), butin(13) and 3,7,4'-trihydroxyflavone(14). Among them, compound 1 was a new compound, while 2 and 3 were new natural products, 6, 8, 9 and 14 were isolated for the first time from Dalbergiae Odoriferae Lignum. Compounds 1-14 were tested for their cytotoxic activity against human hepatoma cell line BEL-7402, human gastric cancer cell line SCG-7901, human lung cancer cell line A549, human chronic myeloid leukemia cell line K562 and HeLa human cervical cancer cellline by MTT method. Compound 1 exhibited significant cytotoxicity with IC_(50) values ranging from 2.85 to 11.62 µg·mL~(-1). In addition, 2, 11 and 12 showed weak cytotoxic activities.

A Biodegradable Polyethylenimine-Based Vector Modified by Trifunctional Peptide R18 for Enhancing Gene Transfection Efficiency In Vivo.

Lack of capacity to cross the nucleus membrane seems to be one of the main reasons for the lower transfection efficiency of gene vectors observed in vivo study than in vitro. To solve this problem, a new non-viral gene vector was designed. First, a degradable polyethylenimine (PEI) derivate was synthesized by crosslinking low-molecular-weight (LMW) PEI with N-octyl-N-quaternary chitosan (OTMCS), and then adopting a designed trifunctional peptide (RGDC-TAT-NLS) with good tumor targeting, cell uptake and nucleus transport capabilities to modify OTMCS-PEI. The new gene vector was termed as OTMCS-PEI-R18 and characterized in terms of its chemical structure and biophysical parameters. Gene transfection efficiency and nucleus transport mechanism of this vector were also evaluated. The polymer showed controlled degradation and remarkable buffer capabilities with the particle size around 100-300 nm and the zeta potential ranged from 5 mV to 40 mV. Agraose gel electrophoresis showed that OTMCS-PEI-R18 could effectively condensed plasmid DNA at a ratio of 1.0. Besides, the polymer was stable in the presence of sodium heparin and could resist digestion by DNase I at a concentration of 63U DNase I/DNA. OTMCS-PEI-R18 also showed much lower cytotoxicity and better transfection rates compared to polymers OTMCS-PEI-R13, OTMCS-PEI and PEI 25 KDa in vitro and in vivo. Furthermore, OTMCS-PEI-R18/DNA complexes could accumulate in the nucleus well soon and not rely on mitosis absolutely due to the newly incorporated ligand peptide NLS with the specific nuclear delivery pathway indicating that the gene delivery system OTMCS-PEI-R18 could reinforce gene transfection efficiency in vivo.

Low-Molecular Weight Polyethylenimine Modified with Pluronic 123 and RGD- or Chimeric RGD-NLS Peptide: Characteristics and Transfection Efficacy of Their Complexes with Plasmid DNA.

To solve the problem of transfection efficiency vs. cytotoxicity and tumor-targeting ability when polyethylenimine (PEI) was used as a nonviral gene delivery vector, new degradable PEI polymers were synthesized via cross-linking low-molecular-weight PEI with Pluronic P123 and then further coupled with a targeting peptide R4 (RGD) and a bifunctional R11 (RGD-NLS), which were termed as P123-PEI-R4 and P123-PEI-R11, respectively. Agarose gel electrophoresis showed that both P123-PEI-R4 and P123-PEI-R11 efficaciously condense plasmid DNA at a polymer-to-pDNA w/w ratio of 3.0 and 0.4, respectively. The polyplexes were stable in the presence of serum and could protect plasmid DNA against DNaseI. They had uniform spherical nanoparticles with appropriate sizes around 100-280 nm and zeta-potentials about +40 mV. Furthermore, in vitro experiments showed that these polyplexes had lower cytotoxicity at any concentration compared with PEI 25 kDa, thus giving promise to high transfection efficiency as compared with another P123-PEI derivate conjugated with trifunctional peptide RGD-TAT-NLS (P123-PEI-R18). More importantly, compared with the other polymers, P123-PEI-R11 showed the highest transfection efficiency with relatively lower cytotoxicity at any concentration, indicating that the new synthetic polymer P123-PEI-R11 could be used as a safe and efficient gene deliver vector.

Coupling of a bifunctional peptide R13 to OTMCS-PEI copolymer as a gene vector increases transfection efficiency and tumor targeting.

BACKGROUND: A degradable polyethylenimine (PEI) derivative coupled to a bifunctional peptide R13 was developed to solve the transfection efficiency versus cytotoxicity and tumor-targeting problems of PEI when used as a gene vector. METHODS: We crossed-linked low molecular weight PEI with N-octyl-N-quaternary chitosan (OTMCS) to synthesize a degradable PEI derivative (OTMCS-PEI), and then used a bifunctional peptide, RGDC-TAT (49-57) called R13 to modify OTMCS-PEI so as to prepare a new gene vector, OTMCS-PEI-R13. This new gene vector was characterized by various physicochemical methods. Its cytotoxicity and gene transfection efficiency were also determined both in vitro and in vivo. RESULTS: The vector showed controlled degradation and excellent buffering capacity. The particle size of the OTMCS-PEI-R13/DNA complexes was around 150-250 nm and the zeta potential ranged from 10 mV to 30 mV. The polymer could protect plasmid DNA from being digested by DNase I at a concentration of 23.5 U DNase I/µg DNA. Further, the polymer was resistant to dissociation induced by 50% fetal bovine serum and 400 µg/mL sodium heparin. Compared with PEI 25 kDa, the OTMCS-PEI-R13/DNA complexes showed higher transfection efficiency both in vitro and in vivo. Further, compared with OTMCS-PEI, distribution of OTMCS-PEI-R13 at tumor sites was markedly enhanced, indicating the tumor-targeting specificity of R13. CONCLUSION: OTMCS-PEI-R13 could be a potential candidate as a safe and efficient gene delivery carrier for gene therapy.