Fresh Washed Microbiota Transplantation Alters Gut Microbiota Metabolites to Ameliorate Sleeping Disorder Symptom of Autistic Children.

Accumulating studies have raised concerns about gut dysbiosis associating autism spectrum disorder (ASD) and its related symptoms. However, the effect of gut microbiota modification on the Chinese ASD population and its underlying mechanism were still elusive. Herein, we enrolled 24 ASD children to perform the first course of fresh washed microbiota transplantation (WMT), 18 patients decided to participate the second course, 13 of which stayed to participate the third course, and there were 8 patients at the fourth course. Then we evaluated the effects of fresh WMT on these patients and their related symptoms. Our results found that the sleeping disorder symptom was positively interrelated to ASD, fresh WMT significantly alleviated ASD and its sleeping disorder and constipation symptoms. In addition, WMT stably and continuously downregulated Bacteroides/Flavonifractor/Parasutterella while upregulated Prevotella_9 to decrease toxic metabolic production and improve detoxification by regulating glycolysis/myo-inositol/D-glucuronide/D-glucarate degradation, L-1,2-propanediol degradation, fatty acid ß-oxidation. Thus, our results suggested that fresh WMT moderated gut microbiome to improve the behavioral and sleeping disorder symptoms of ASD via decrease toxic metabolic production and improve detoxification. Which thus provides a promising gut ecological strategy for ASD children and its related symptoms treatments.

Potential prognostic and immunotherapeutic value of calponin 1: A pan-cancer analysis.

Background: Emerging evidence has suggested a pro-oncogenic role of calponin 1 (CNN1) in the initiation of a variety of cancers. Despite this, CNN1 remains unknown in terms of its effects and mechanisms on angiogenesis, prognosis, and immunology in cancer. Materials and Methods: The expression of CNN1 was extracted and analyzed using the TIMER, UALCAN, and GEPIA databases. Meanwhile, we analyzed the diagnostic value of CNN1 by using PrognoScan and Kaplan-Meier plots. To elucidate the value of CNN1 in immunotherapy, we used the TIMER 2.0 database, TISIDB database, and Sangerbox database. Gene set enrichment analysis (GSEA) was used to analyze the expression pattern and bio-progression of CNN1 and the vascular endothelium growth factor (VEGF) in cancer. The expressions of CNN1 and VEGF in gastric cancer were confirmed using immunohistochemistry. We used Cox regression analysis to investigate the association between pathological characteristics, clinical prognosis, and CNN1 and VEGF expressions in patients with gastric cancer. Results: CNN1 expression was higher in normal tissues than it was in tumor tissues of most types of cancers. However, the expression level rebounds during the development of tumors. High levels of CNN1 indicate a poor prognosis for 11 tumors, which include stomach adenocarcinoma (STAD). There is a relationship between CNN1 and tumor-infiltrating lymphocytes (TILs), and the marker genes NRP1 and TNFRSF14 of TILs are significantly related to CNN1 expression in gastric cancers. The GSEA results confirmed the lower expression of CNN1 in tumors when compared to normal tissues. However, CNN1 again showed an increasing trend during tumor development. In addition, the results also suggest that CNN1 is involved in angiogenesis. The immunohistochemistry results validated the GSEA result (take gastric cancer as an example). Cox analysis suggested that high CNN1 expression and high VEGF expression are closely associated with poor clinical prognosis. Conclusion: Our study has shown that CNN1 expression is aberrantly elevated in various cancers and positively correlates with angiogenesis and the immune checkpoint, contributing to cancer progression and poor prognosis. These results suggest that CNN1 could serve as a promising candidate for pan-cancer immunotherapy.

Recent developments in mesoporous polydopamine-derived nanoplatforms for cancer theranostics.

Polydopamine (PDA), which is derived from marine mussels, has excellent potential in early diagnosis of diseases and targeted drug delivery owing to its good biocompatibility, biodegradability, and photothermal conversion. However, when used as a solid nanoparticle, the application of traditional PDA is restricted because of the low drug-loading and encapsulation efficiencies of hydrophobic drugs. Nevertheless, the emergence of mesoporous materials broaden our horizon. Mesoporous polydopamine (MPDA) has the characteristics of a porous structure, simple preparation process, low cost, high specific surface area, high light-to-heat conversion efficiency, and excellent biocompatibility, and therefore has gained considerable interest. This review provides an overview of the preparation methods and the latest applications of MPDA-based nanodrug delivery systems (chemotherapy combined with radiotherapy, photothermal therapy combined with chemotherapy, photothermal therapy combined with immunotherapy, photothermal therapy combined with photodynamic/chemodynamic therapy, and cancer theranostics). This review is expected to shed light on the multi-strategy antitumor therapy applications of MPDA-based nanodrug delivery systems.

The Potential of Drug Delivery Nanosystems for Sepsis Treatment.

Sepsis is a major immune response disorder caused by infection, with very high incidence and mortality rates. In the clinic, sepsis and its complications are mainly controlled and treated with antibiotics, anti-inflammatory, and antioxidant drugs. However, these treatments have some shortcomings, such as rapid metabolism and severe side effects. The emergence of drug delivery nanosystems can significantly improve tissue permeability, prolong drugs' circulation time, and reduce side effects. In this paper, we reviewed recent drug delivery nanosystems designed for sepsis treatment based on their mechanisms (anti-bacterial, anti-inflammatory, and antioxidant). Although great progress has been made recently, clinical practice transformation is still very difficult. Therefore, we also discussed key obstacles, including tissue distribution, overcoming bacterial resistance, and single treatment modes. Finally, a rigorous optimization of drug delivery nanosystems is expected to present great potential for sepsis therapy.

Preparation of environmentally friendly acrylic pressure-sensitive adhesives by bulk photopolymerization and their performance.

Polyacrylic pressure-sensitive adhesives (PSAs) based on butyl acrylate (BA), 2-hydroxyethyl acrylate (HEA), and acrylic acid (AA) were prepared by a bulk polymerization process triggered by a radical photoinitiator under UV irradiation and UV-crosslinking. 1,6-Hexanediol diacrylate (HDDA) with difunctional groups was introduced into the PSAs to modify semi-interpenetrating network structures. The effect of HDDA content on the pressure-sensitive performance was comprehensively tested. The viscosity of the prepolymer was measured by a rotational viscometer. Prepolymers obtained by a photoinduced process and UV crosslinking process were confirmed via Fourier transform infrared spectroscopy (FTIR). All double bonds participated in the copolymerization without any remaining monomers, which reflected the concept of green environmental protection. Gel content in the crosslinked portion was examined by Soxhlet extraction, whilst the soluble molecular weight of PSAs was characterized by gel permeation chromatography (GPC). The viscoelastic properties of polymer films were determined by dynamic mechanical analysis (DMA). The T g value and storage modulus (G') of the PSAs were enhanced with the addition of HDDA. Moreover, three fundamental adhesive properties, i.e. loop tack force, peel force and shear strength of PSAs, were measured. The results showed that UV crosslinking technology achieved a good balance of the three forces with excellent pressure-sensitive properties.

Biochemical characterization and structural analysis of ulvan lyase from marine Alteromonas sp. reveals the basis for its salt tolerance.

Marine macroalgae have gained considerable attention as renewable biomass sources. Ulvan is a water-soluble anionic polysaccharide, and its depolymerization into fermentable monosaccharides has great potential for the production of bioethanol or high-value food additives. Ulvan lyase from Alteromonas sp. (AsPL) utilizes a ß-elimination mechanism to cleave the glycosidic bond between rhamnose 3-sulfate and glucuronic acid, forming an unsaturated uronic acid at the non-reducing end. AsPL was active in the temperature range of 30-50 °C and pH values ranging from 7.5 to 9.5. Furthermore, AsPL was found to be halophilic, showing high activity and stability in the presence of up to 2.5 M NaCl. The apparent Km and kcat values of AsPL are 3.19 ±â€¯0.37 mg mL-1 and 4.19 ±â€¯0.21 s-1, respectively. Crystal structure analysis revealed that AsPL adopts a ß-propeller fold with four anti-parallel ß-strands in each of the seven propeller blades. The acid residues at the protein surface and two Ca2+ coordination sites contribute to its salt tolerance. The research on ulvan lyase has potential commercial value in the utilization of algal resources for biofuel production to relieve the environmental burden of petrochemicals.

Engineering a thermostable version of D-allulose 3-epimerase from Rhodopirellula baltica via site-directed mutagenesis based on B-factors analysis.

D-allulose has received increasing attention due to its excellent physiological properties and commercial potential. The D-allulose 3-epimerase from Rhodopirellula baltica (RbDAEase) catalyzes the conversion of D-fructose to D-allulose. However, its poor thermostability has hampered its industrial application. Site-directed mutagenesis based on homologous structures in which the residuals on high flexible regions were substituted according to B-factors analysis, is an effective way to improve the thermostability and robustness of an enzyme. RbDAEase showed substrate specificity toward D-allulose with a Km of 58.57 mM and kcat of 1849.43 min-1. It showed a melting temperature (Tm) of 45.7 °C and half-life (t1/2) of 52.3 min at pH 8.0, 60 °C with 1 mM Mn2+. The Site-directed mutation L144 F strengthened the thermostability to a Δt1/2 of 50.4 min, ΔTm of 12.6 °C, and ΔT5060 of 22 °C. It also improved the conversion rate to 28.6%. Structural analysis reveals that a new hydrophobic interaction was formed by the mutation. Thus, site-directed mutagenesis based on B-factors analysis would be an efficient strategy to enhance the thermostability of designed ketose 3-epimerases.

Efficient Biosynthesis of High-Value Succinic Acid and 5-Hydroxyleucine Using a Multienzyme Cascade and Whole-Cell Catalysis.

Succinic acid (SA) is applied in the food, chemical, and pharmaceutical industries. 5-Hydroxyleucine (5-HLeu) is a promising precursor for the biosynthesis of antituberculosis drugs. Here, we designed a promising synthetic route for the simultaneous production of SA and 5-HLeu by combining l-leucine dioxygenase (NpLDO), l-glutamate oxidase (LGOX), and catalase (CAT). Two bioconversion systems: "a multienzyme cascade catalysis in vitro" (MECCS) and "whole-cell catalysis system" (WCCS) were constructed. A high-activity NpLDO mutant was screened by error-prone polymerase chain reaction (PCR) and showed 6.1-fold improvement of catalytic activity. After optimization of reaction conditions, MECSS yielded 3.15 g/L SA and 3.92 g/L 5-HLeu, while the production of SA and 5-HLeu by the most effective WCSS reached 15.12 and 18.83 g/L, respectively. This is the first attempt to use ferrous iron/α-ketoglutarate-dependent dioxygenases for the simultaneous production of SA and hydroxy-amino-acid. This research provides a tool for industrial production of food of high-value products from low-cost raw materials.

Soluble expression, purification and biochemical characterization of a C-7 cholesterol dehydrogenase from Drosophila melanogaster.

The C-7 cholesterol dehydrogenase (NVD), which converts cholesterol to 7-dehydrocholesterol (7-DHC) by 7,8-dehydrogenation, plays a pivotal role in the metabolism of cholesterol and steroid intermediates. The NVD protein was successfully expressed in insect Sf9 cells. To reduce the production cost for industrial application, the NVD gene was cloned into E. coli BL21(DE3). However, the His-tagged NVD protein showed poor binding to Ni-NTA resin, mainly due to the formation of inclusion bodies. Consequently, the expression and solubility of NVD were optimized by respectively fusing it with maltose-binding protein (MBP), glutathione S-transferase (GST), and the nonspecific DNA binding protein from Sulfolobus solfataricus (Sso7d) as solubility tags. The NVD fusion with MBP at the N-terminus and His-tag at the C-terminus achieved a high yield of the soluble enzyme. It was further purified by ion-exchange chromatography with 95.4% purity and with a 10.4% yield. The product 7-DHC, which is produced in a reaction catalyzed by NVD and ferredoxin reductase KshB, was initially identified by GC-MS. An analysis of the amino acid sequence alignment revealed a distinct Rieske-type iron-sulfur cluster and non-heme Fe2+-binding domain, which are evolutionarily conserved among NVD family enzymes.

Highly Integrated Nanoplatform Based on an E-Selectin-Targeting Strategy for Metastatic Breast Cancer Treatment.

Therapeutic goals for metastatic breast cancer, including shrinkage of established metastasis and suppression of movement of tumor cells, are often hard to achieve and remain the main obstacles restricting the antimetastatic efficacy of targeted drug delivery systems (TDDSs). Herein, we proposed an E-selectin-targeting nanoplatform for the systemic treatment of metastatic breast cancer. Versatile functions, including killing the circulating tumor cells, shrinking the established lesions, as well as inhibiting the movement of tumor cells, were integrated into doxorubicin-loaded sialic acid-dextran-octadecanoic acid (SDO) micelles (SDD). The prepared SDD micelles could not only inhibit lung and liver metastasis in the orthotopic 4T1 tumors model, but also decrease the metastatic lesions in the metastatic 4T1 cell model, resulting in 27.33% reduced number of metastatic nodules when compared to those without sialic acid modification. It was found that the good antimetastatic effect of SDD was only partially attributed to its ability on removing metastatic cells and metastases. Most importantly, the blank SDO micelles left in the lesion could further inhibit the cell migration and cell-cell binding. These results suggest that SA-driven TDDS has the potential for specific targeting and effective treatment of cancer metastasis.

A dual deformable liposomal ointment functionalized with retinoic acid and epidermal growth factor for enhanced burn wound healing therapy.

An ointment containing retinoic acid deformable liposomes (TRA DLs) and epidermal growth factor cationic deformable liposomes (EGF CDLs) was prepared for the treatment of deep partial-thickness burns. The characterization tests confirmed both liposomes featured small particle sizes, high drug entrapment efficiencies and sustained drug release behavior. Compared with the free drug, TRA DLs and EGF CDLs exhibited superior skin permeation and remarkably increased drug deposition by 2.9 and 18.8 folds, respectively. Results on HaCaT cells indicated the combined application of two liposomes exerted a synergistic effect and prominently promoted cell proliferation and migration. Application of the dual liposomal ointment on a deep partial-thickness burn model stimulated wound closure (p < 0.001), promoted skin appendage formation and increased collagen production, thus improving healing quality. Finally, it was demonstrated that TRA significantly up-regulated the expression of EGFR and HB-EGF to enhance the therapeutic effect of EGF. Therefore, the dual liposomal ointment is a promising topical therapeutic for burn treatment.

Redesign of a novel D-allulose 3-epimerase from Staphylococcus aureus for thermostability and efficient biocatalytic production of D-allulose.

BACKGROUND: A novel D-allulose 3-epimerase from Staphylococcus aureus (SaDAE) has been screened as a D-allulose 3-epimerase family enzyme based on its high specificity for D-allulose. It usually converts both D-fructose and D-tagatose to respectively D-allulose and D-sorbose. We targeted potential biocatalysts for the large-scale industrial production of rare sugars. RESULTS: SaDAE showed a high activity on D-allulose with an affinity of 41.5 mM and catalytic efficiency of 1.1 s-1 mM-1. Four residues, Glu146, Asp179, Gln205, and Glu240, constitute the catalytic tetrad of SaDAE. Glu146 and Glu240 formed unique interactions with substrates based on the structural model analysis. The redesigned SaDAE_V105A showed an improvement of relative activity toward D-fructose of 68%. The conversion rate of SaDAE_V105A reached 38.9% after 6 h. The triple mutant S191D/M193E/S213C showed higher thermostability than the wild-type enzyme, exhibiting a 50% loss of activity after incubation for 60 min at 74.2 °C compared with 67 °C for the wild type. CONCLUSIONS: We redesigned SaDAE for thermostability and biocatalytic production of D-allulose. The research will aid the development of industrial biocatalysts for D-allulose.

Increasing HER2 α2,6 sialylation facilitates gastric cancer progression and resistance via the Akt and ERK pathways.

Upregulated ß-galactoside α2,6-sialyltransferase I (ST6Gal-I) expression reportedly occurs in many cancers and is correlated with metastasis and poor prognosis. However, the mechanisms by which ST6Gal­I facilitates gastric cancer progression remain poorly understood. Trastuzumab is exclusively used in human epidermal growth factor receptor 2 (HER2)+ gastric cancers; however, most advanced HER2+ gastric cancers develop trastuzumab resistance. Herein, we identified HER2 as an ST6Gal­I substrate and showed that HER2 α2,6 sialylation confers protection against trastuzumab­mediated apoptosis. SGC7901 cancer cell models in which ST6Gal­I was overexpressed or knocked down were constructed, revealing that ST6Gal­I overexpression induced high HER2 sialylation levels and increased cell viability and invasion compared to those in the vector cell line under serum starvation; ST6Gal­I knockdown had the opposite effects. ST6Gal­I overexpression also potentiated cell cycle arrest in the G2/S phase to reduce drug sensitivity. In addition, FACS analysis revealed that high ST6Gal­I levels increased resistance to trastuzumab­induced apoptosis, accompanied by decreased caspase­3 levels. However, the ST6Gal­I knockdown cell line revealed increased caspase­3 levels and evident apoptosis compared with those in the vector cell line. Although ST6Gal­I overexpression increased HER2 sialylation, corresponding to decreased HER2 phosphorylation, high α2,6­sialylation enhanced Akt and ERK phosphorylation levels compared to those in the vector cell line; ST6Gal­I knockdown had the opposite effects. Collectively, these results implicated a functional role of ST6Gal­I in promoting tumor cell progression and trastuzumab resistance.

Sialic Acid-Functionalized pH-Triggered Micelles for Enhanced Tumor Tissue Accumulation and Active Cellular Internalization of Orthotopic Hepatocarcinoma.

Both targeted and stimuli-sensitive drug-delivery systems (DDSs) have been developed to augment antitumor effects. However, lack of knowledge regarding tumor tissue targeting and different effects of the stimuli-sensitive DDSs in orthotropic and ectopic tumors have impeded further advances in their clinical applications. Herein, we first reported a pH-triggered micelle with sialic acid (SA)-driven targeting ability (SA-poly(ethylene glycol)-hydrazone linker-doxorubicin (DOX), SPD). The SPD micelles encapsulated with DOX (SPDD) showed sustained drug release over 48 h in response to the pH gradient in vivo, slow under physical conditions and accelerated in the acid tumor microenvironment. In addition, the SPD micelles showed 2.3-fold higher accumulation in tumors after 48 h compared to the micelles lacking the SA moiety. The overexpression of E-selectin on the inflammatory vascular endothelial cells surrounding the tumors increased the accumulation of SPD micelles in tumor tissues, whereas that on the tumor cells increased the internalization of micelles. Consequently, SPDD micelles exerted remarkable antitumor effects in both orthotopic and ectopic models. Application of SPDD micelles in the in situ model reduced the tumor volume (77.57 mm3 vs 62.13 mm3) and metastasis after treatment for 25 days. These results suggest that SA-driven targeted DDS with a pH-responsive switch has the potential to treat hepatocarcinoma effectively both ectopically and orthotopically.

Sialic-Acid-Anchored Micelles: A Hierarchical Targeting Device for Enhanced Tumor Tissue Accumulation and Cellular Internalization.

Targeted drug delivery systems (TDDS) have attracted wide attention for their reduced drug side effects and improved antitumor efficacy in comparison with traditional preparations. While targeting moieties in existing TDDS have principally focused on recognition of receptors on the surface of tumor cells, accumulation into tumor tissue only could be performed by enhanced permeability and retention effects and active transportation into tumor cells. Doxorubicin (DOX)-loaded sialic acid-dextran (Dex)-octadecanoic acid (OA) micelles (SA-Dex-OA/DOX) were designed for targeting hepatocellular carcinoma effectively. The synthesized conjugates could self-aggregate to form micelles with a critical micelle concentration of 27.6 µg·mL-1 and diameter of 54.53 ± 3.23 nm. SA-Dex-OA micelles incorporated with 4.36% DOX-loading content could prolong in vitro drug release to 96 h with 80% of final release. Cellular transportation studies revealed that SA-Dex-OA micelles mediated more efficient DOX delivery into Bel-7402 cells than those without SA modification. In vivo biodistribution testing demonstrated that SA-Dex-OA/ICG micelles showed 3.05-fold higher accumulation into Bel-7402 tumors. The recognition of overexpressed E-selectin in inflammatory tumor vascular endothelial cells led to a large accumulation of SA-Dex-OA/ICG micelles into tumor tissue, and the E-selectin upregulated on the surface of tumor cells contributed to active cellular transportation into tumor cells. Accordingly, SA-Dex-OA/DOX exhibited prior suppression of Bel-7402 tumor growth greater than that of Dex-OA/DOX micelles and free DOX (the tumor inhibition: 79.2% vs 61.0 and 51.3%). These results suggest that SA-functionalized micelles with dual targeting properties have high potential for liver cancer therapy.

Visual targeted therapy of hepatic cancer using homing peptide modified calcium phosphate nanoparticles loading doxorubicin guided by T1 weighted MRI.

Effective treatment and real-time monitoring of hepatic cancer are essential. A multifunctional calcium phosphate nanoparticles loading chemotherapeutic agent doxorubicin and magnetic resonance imaging contrast agent diethylenetriaminepentaacetic acid gadolinium (A54-CaP/Gd-DTPA/DOX) was developed for visual targeted therapy of hepatic cancer via T1-weighted MRI in real-time. A54-CaP/Gd-DTPA/DOX exhibited a higher longitudinal relaxivity (6.02 mM-1 s-1) than commercial MR contrast agent Gd-DTPA (3.3765 mM-1 s-1). The DOX release from the nanoparticles exhibited a pH dependent behavior. The cellular uptake results showed that the internalization of A54-CaP/Gd-DTPA/DOX into BEL-7402 cells was1.9-fold faster than that of HepG2 cells via A54 binding. In vivo experiments presented that A54-CaP/Gd-DTPA/DOX had higher distribution and longer retention time in tumor tissue than CaP/Gd-DTPA/DOX and free DOX, and also displayed great antitumor efficacy (95.38% tumor inhibition rate) and lower toxicity. Furthermore, the Gd-DTPA entrapped in the nanoparticles could provide T1-weighted MRI for real-time monitoring the progress of tumor treatment.