Application of PHA surface binding proteins of alkali-tolerant Bacillus as surfactants.

Amphiphilic protein has lipophilic and hydrophilic domains, displaying the potential for development as a biosurfactant. The polyhydroxyalkanoate (PHA) surface binding protein derived from Bacillus is a type of protein that has not been studied for its emulsifying properties. In this study, PHA granule-associated protein (PhaP), PHA regulatory protein (PhaQ), and PHA synthase subunit (PhaR) derived from an alkali-tolerant PHA-producing Bacillus cereus HBL-AI were found and heterologously expressed in E. coli and purified to investigate their application as biosurfactants. It showed that the emulsification ability and stability of three amphiphilic proteins were higher than those of widely used chemical surfactants in diesel oil, vegetable oil, and lubricating oil. In particular, the PhaQ protein studied for the first time can form a stable emulsion layer in vegetable oil at a lower concentration (50 µg/mL), which greatly reduced the amount of protein used in emulsification. This clearly demonstrated that the PHA-binding protein of HBL-AI can be well applied as an environmentally friendly biosurfactants.

Traditional Chinese medicine residue enzymatic hydrolysates for production of polyhydroxyalkanoate by newly isolated Bacillus altitudinis.

Traditional Chinese medicine residue (TCMR) was utilized as an inexpensive carbon source for the production of poly(3-hydroxybutyrate) (PHB) using the newly isolated Bacillus altitudinis HBU-SI7. The results showed that Yu Ping Feng TCMR could be directly hydrolysed by cellulase to obtain a high proportion of glucose (99 % of total sugar) without pretreatment, achieving an enzymatic hydrolysis rate of up to 89.2 %. B. altitudinis could grow and produce PHB when using enzymatically hydrolysed TCMR in a 5-L fermenter. After 20 h of fermentation, the maximum concentration of PHB was 11.2 g/L, and the highest cell dry weight (CDW) was 15.4 g/L, with 72.7 % of the PHB fraction in CDW. Moreover, this strain could utilize enzymatic hydrolysates from various herbal formulas to produce high levels of PHB. This novel approach aims to accumulate PHB from TCMR hydrolysates, offering an effective and environmentally friendly method to reduce production costs and achieve mass production.

Targeted PHA Microsphere-Loaded Triple-Drug System with Sustained Drug Release for Synergistic Chemotherapy and Gene Therapy.

The combination of paclitaxel (PTX) with other chemotherapy drugs (e.g., gemcitabine, GEM) or genetic drugs (e.g., siRNA) has been shown to enhance therapeutic efficacy against tumors, reduce individual drug dosages, and prevent drug resistance associated with single-drug treatments. However, the varying solubility of chemotherapy drugs and genetic drugs presents a challenge in co-delivering these agents. In this study, nanoparticles loaded with PTX were prepared using the biodegradable polymer material poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx). These nanoparticles were surface-modified with target proteins (Affibody molecules) and RALA cationic peptides to create core-shell structured microspheres with targeted and cationic functionalization. A three-drug co-delivery system (PTX@PHBHHx-ARP/siRNAGEM) were developed by electrostatically adsorbing siRNA chains containing GEM onto the microsphere surface. The encapsulation efficiency of PTX in the nanodrug was found to be 81.02%, with a drug loading of 5.09%. The chemogene adsorption capacity of siRNAGEM was determined to be 97.3%. Morphological and size characterization of the nanodrug revealed that PTX@PHBHHx-ARP/siRNAGEM is a rough-surfaced microsphere with a particle size of approximately 150 nm. This nanodrug exhibited targeting capabilities toward BT474 cells with HER2 overexpression while showing limited targeting ability toward MCF-7 cells with low HER2 expression. Results from the MTT assay demonstrated that PTX@PHBHHx-ARP/siRNAGEM exhibits high cytotoxicity and excellent combination therapy efficacy compared to physically mixed PTX/GEM/siRNA. Additionally, Western blot analysis confirmed that siRNA-mediated reduction of Bcl-2 expression significantly enhanced cell apoptosis mediated by PTX or GEM in tumor cells, thereby increasing cell sensitivity to PTX and GEM. This study presents a novel targeted nanosystem for the co-delivery of chemotherapy drugs and genetic drugs.

Preparation of polyhydroxyalkanoate-based magnetic microspheres for carbonyl reductase purification and immobilization.

A polyhydroxyalkanoate (PHA) magnetic microsphere was designed for one-step purification and immobilization of a novel carbonyl reductase (RLSR5) from recombinant Escherichia coli lysate. The hydrophobic core of this microsphere was composed of a highly biocompatible polymer, poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx), in which magnetic Fe3O4 particles were embedded during solvent evaporation. The hydrophilic shell of the fusion protein formed by PHA particle-binding protein (PhaP) and RLSR5 (PR) was expressed in recombinant E. coli. The magnetic core of Fe3O4@PHBHHx directly purified the hydrophilic shell from the E. coli lysate, and the two self-assembled to form Fe3O4@PHBHHx-PR through hydrophobic and hydrophilic interactions, eliminating the separation of the fusion protein. The microstructure, magnetic properties, morphology, size, and dispersion of Fe3O4@PHBHHx-PR were investigated by XRD, VSM, SEM, TEM, elemental mapping and DLS. It was found that Fe3O4@PHBHHx-PR correctly assembled, with a well dispersed spherical structure at the nanoscale and superparamagnetism properties. The amount of RLSR5 immobilized on PHA microspheres reached 121.9 mg/g. The Fe3O4@PHBHHx-PR was employed to synthesize (R)-tolvaptan with 99 % enantiomeric excess and 97 % bioconversion efficiency, and the catalyst maintained 78.6 % activity after 10 recovery cycles. These PHA magnetic microspheres are versatile carriers for enzyme immobilization and demonstrate improved stability and reusability of the free enzyme.

Heart failure with preserved ejection fraction and non-alcoholic fatty liver disease: new insights from bioinformatics.

AIMS: Heart failure with preserved ejection fraction (HFpEF) and non-alcoholic fatty liver disease (NAFLD) are related conditions with an increasing incidence. The mechanism of their relationship remains undefined. Here, we aimed to explore the potential mechanisms, diagnostic markers, and therapeutic options for HFpEF and NAFLD. METHODS AND RESULTS: HFpEF and NAFLD datasets were downloaded from the Gene Expression Omnibus (GEO) database. Common differentially expressed genes (DEGs) were screened for functional annotation. A protein-protein interaction network was constructed based on the STRING database, and hub genes were analysed using GeneMANIA annotation. ImmuCellAI (Immune Cell Abundance Identifier) was employed for analysis of immune infiltration. We also used validation datasets to validate the expression levels of hub genes and the correlation of immune cells. To screen for diagnostic biomarkers, we employed the least absolute shrinkage and selection operator and support vector machine-recursive feature elimination. Drug signature database was used to predict potential therapeutic drugs. Our analyses identified a total of 33 DEGs. Inflammation and immune infiltration played important roles in the development of both diseases. The data showed a close relationship between chemokine signalling pathway, cytokine-cytokine receptor interaction, calcium signalling pathway, neuroactive ligand-receptor interaction, osteoclast differentiation, and cyclic guanosine monophosphate-protein kinase G signalling pathway. We demonstrated that PRF1 (perforin 1) and IL2RB (interleukin-2 receptor subunit beta) proteins were perturbed by the diseases and may be the hub genes. The analysis showed that miR-375 may be a potential diagnostic marker for both diseases. Our drug prediction analysis showed that bosentan, eldecalcitol, ramipril, and probucol could be potential therapeutic options for the diseases. CONCLUSIONS: Our findings revealed common pathogenesis, diagnostic markers, and therapeutic agents for HFpEF and NAFLD. There is need for further experimental studies to validate our findings.

A bibliometric analysis of myocardial ischemia/reperfusion injury from 2000 to 2023.

Background: Myocardial ischemia/reperfusion injury (MIRI) refers to the more severe damage that occurs in the previously ischemic myocardium after a short-term interruption of myocardial blood supply followed by restoration of blood flow within a certain period of time. MIRI has become a major challenge affecting the therapeutic efficacy of cardiovascular surgery. Methods: A scientific literature search on MIRI-related papers published from 2000 to 2023 in the Web of Science Core Collection database was conducted. VOSviewer was used for bibliometric analysis to understand the scientific development and research hotspots in this field. Results: A total of 5,595 papers from 81 countries/regions, 3,840 research institutions, and 26,202 authors were included. China published the most papers, but the United States had the most significant influence. Harvard University was the leading research institution, and influential authors included Lefer David J., Hausenloy Derek J., Yellon Derek M., and others. All keywords can be divided into four different directions: risk factors, poor prognosis, mechanisms and cardioprotection. Conclusion: Research on MIRI is flourishing. It is necessary to conduct an in-depth investigation of the interaction between different mechanisms and multi-target therapy will be the focus and hotspot of MIRI research in the future.

Amphiphilic Cationic Peptide-Coated PHA Nanosphere as an Efficient Vector for Multiple-Drug Delivery.

Amphiphilic core-shell (ACS) nanoparticles are gaining increasing research interest for multi-drug delivery in cancer therapy. In this work, a new cationic peptide-coated PHA nanosphere was prepared by self-assembly of a hydrophobic core of biodegradable poly (3-hydroxybutyrate-co-3-hydroxyhexanoate) (PHBHHx) and a hydrophilic shell of fusion proteins of PHA granule-associated protein (PhaP) and cationic peptide RALA through a strong hydrophobic effect. The hydrophobic drug curcumin (Cur) was encapsulated in PHBHHx nanoparticles. The chemotherapy drug 5-fluorouracil (5-FU) was administered in the form of its metabolite oligomeric 5-fluorodeoxyuridine (FUdR). Fifteen consecutive FUdR (FUdR15S) were adsorbed on the surface of PHBHHx nanoparticles by electrostatic interaction with RALA to form Cur@PHBX-PR/FUdR15S. Such amphiphilic cationic nanospheres had 88.3% EE of Cur and the drug loading of Cur and FUdR were 7.8% and 12.1%. The dual-drug-loaded nanospheres showed a time-differential release of Cur and FUdR. In addition, Cur@PHBX-PR/FUdR15S exhibited excellent anticancer activity and played a vital role in promoting the synergistic effect of FUdR and Cur in gastric cancer cells. The exploration of antitumor mechanisms demonstrated that Cur improved the activity of apoptosis-related proteins and cancer cells sensitized to FUdR. This amphiphilic core-shell system can serve as a general platform for sequential delivery of multiple drugs to treat several cancer cells.

Affibody Modified G-quadruplex DNA Micelles Incorporating Polymeric 5-Fluorodeoxyuridine for Targeted Delivery of Curcumin to Enhance Synergetic Therapy of HER2 Positive Gastric Cancer.

Combination chemotherapy is emerging as an important strategy for cancer treatment with decreased side effects. However, chemotherapeutic drugs with different solubility are not easy to realize co-delivery in traditional nanocarriers. Herein, an affibody modified G-quadruplex DNA micellar prodrug (affi-F/GQs) of hydrophilic 5-fluorodeoxyuridine (FUdR) by integrating polymeric FUdRs into DNA strands is developed for the first time. To achieve synergistic efficacy with hydrophobic drugs, curcumin (Cur) is co-loaded into affi-F/GQs micelles to prepare the dual drug-loaded DNA micelles (Cur@affi-F/GQs), in which affibody is employed as a targeting moiety to facilitate HER2 receptor-mediated uptake. Cur@affi-F/GQs have a small size of approximately 130 nm and exhibit excellent stability. The system co-delivers FUdR and Cur in a ratiometric manner, and the drug loading rates are 21.1% and 5.6%, respectively. Compared with the physical combination of FUdR and Cur, Cur@affi-F/GQs show higher cytotoxicity and greater synergistic effect on HER2 positive gastric cancer N87 cells. Surprisingly, Cur@affi-F/GQs significantly enhance the expression and activity of apoptosis-associated proteins in Bcl-2/Bax-caspase 8, 9-caspase 3 apoptotic pathway, which is the main factor in the death of tumor cells induced by FUdR. Overall, this nanoencapsulation is a promising candidate for the targeted co-delivery of drugs with significant differences in solubility.

Affibody-Conjugated RALA Polymers Delivering Oligomeric 5-Fluorodeoxyuridine for Targeted Therapy of HER2 Overexpressing Gastric Cancer.

Affibody-conjugated RALA (affi-RA) are designed for delivering oligomeric 5-fluorodeoxyuridine (FUdR, metabolite of 5-FU) strand to raise the selectivity of 5-fluorouracil (5-FU), decrease its toxicity and improve its suboptimal therapeutic efficacy. The nanodrugs, FUdR@affi-RA, are spontaneously assembled by electrostatic interaction between positively charged affi-RA and negatively charged FUdR15 -strands (15 consecutive FUdR). FUdR@affi-RA exhibits excellent stability under simulated physiological conditions. Compared with free FUdR, FUdR@affi-RA shows excellent targeting and higher cytotoxicity in human epidermal growth factor receptor 2 (HER2) overexpressing gastric cancer N87 cells. Moreover, the anticancer mechanism studies reveal that FUdR@affi-RA enhances the expression and activity of apoptosis-associated proteins in the Bcl-2/Bax-caspase 8,9-caspase 3 apoptotic pathway induced by FUdR. This study indicates that the fusion vector, affi-RA, presents a promising delivery system platform for nucleoside analogue drugs and provides a new strategy for the development of therapeutics of cancer treatment.