Rapid diagnosis of largemouth bass ranavirus in fish samples using the loop-mediated isothermal amplification method.

Largemouth bass ranavirus (LMBV) has been recognized as the causative pathogen responsible for infectious skin ulcerative syndrome in cultured largemouth bass in China. A fast and simple LMBV detection method is urgently needed. Here, a loop-mediated isothermal amplification (LAMP) assay was established for the detection of this virus using primers targeting the major capsid protein gene of LMBV. The amplification conditions were optimized; the assay was specific for the diagnosis of LMBV, as there was no cross-reactivity with other four Iridoviridae viruses (large yellow croaker iridovirus, Singapore grouper iridovirus, tiger frog virus, and soft-shelled turtle iridovirus), grass carp reovirus, white spot syndrome virus, or healthy largemouth bass. The sensitivity of the LAMP assay was found to be 8.55 × 101 copies/µL of LMBV DNA, which was 10-fold higher than that of the conventional PCR. Application of the LAMP assay was evaluated using 10 clinical samples, and the results indicated the reliability of the test as a rapid, field diagnostic tool for LMBV detection. Thus, the simplicity and nearly instrument-free LAMP method provides an alternative for rapid and sensitive detection of LMBV and has great potential for early diagnosis of LMBV infection in the farm.

Rapid UV Photo-Cross-Linking of α-Lactalbumin Hydrogel Biomaterial To Enable Wound Healing.

Although both the function and biocompatibility of protein-based biomaterials are better than those of synthetic materials, their usage as medical material is currently limited by their high costs, low yield, and low batch-to-batch reproducibility. In this article, we show how α-lactalbumin (α-LA), rich in tryptophan, was used to produce a novel type of naturally occurring, protein-based biomaterial suitable for wound dressing. To create a photo-cross-linkable polymer, α-LA was methacrylated at a 100-g batch scale with >95% conversion and 90% yield. α-LAMA was further processed using photo-cross-linking-based advanced processing techniques such as microfluidics and 3D printing to create injectable hydrogels, monodispersed microspheres, and patterned scaffolds. The obtained α-LAMA hydrogels show promising biocompatibility and degradability during in vivo testing. Additionally, the α-LAMA hydrogel can accelerate post-traumatic wound healing and promote new tissue regeneration. In conclusion, cheap and safe α-LAMA-based biomaterials could be produced, and they have a beneficial effect on wound healing. As a result, there may arise a potential partnership between the dairy industry and the development of pharmaceuticals.

Copper-loaded Milk-Protein Derived Microgel Preserves Cardiac Metabolic Homeostasis After Myocardial Infarction.

Myocardial Infarction (MI) is a leading cause of death worldwide. Metabolic modulation is a promising therapeutic approach to prevent adverse remodeling after MI. However, whether material-derived cues can treat MI through metabolic regulation is mainly unexplored. Herein, a Cu2+ loaded casein microgel (CuCMG) aiming to rescue the pathological intramyocardial metabolism for MI amelioration is developed. Cu2+ is an important ion factor involved in metabolic pathways, and intracardiac copper drain is observed after MI. It is thus speculated that intramyocardial supplementation of Cu2+ can rescue myocardial metabolism. Casein, a milk-derived protein, is screened out as Cu2+ carrier through molecular-docking based on Cu2+ loading capacity and accessibility. CuCMGs notably attenuate MI-induced cardiac dysfunction and maladaptive remodeling, accompanied by increased angiogenesis. The results from unbiased transcriptome profiling and oxidative phosphorylation analyses support the hypothesis that CuCMG prominently rescued the metabolic homeostasis of myocardium after MI. These findings enhance the understanding of the design and application of metabolic-modulating biomaterials for ischemic cardiomyopathy therapy.

Supplementing maternal diet with milk oligosaccharides and probiotics helps develop the immune system and intestinal flora of offsprings.

Intestinal flora is very important for improving the development of the immune system in newborns. Maternal diet during pregnancy and lactation is one of the key factors affecting the growth and development of offspring. The objective of the present study was to examine whether supplementation of maternal diet with milk oligosaccharides and Bifidobacterium could influence the development of the intestinal flora and immune system of neonatal mice. In total, 30 pregnant Institute of Cancer Research (ICR) mice were randomly divided into six groups: a control group (basal diet) and five intervention groups (basal diet supplemented with different doses of 2'-fucosyllactose [2'-FL] and Bifidobacterium Bb12) during the pregnancy period. All female mice were monitored for physical health during gavage. After delivery, the number of mice in each litter, any deformity, and the development of the offspring were recorded. The spleen, blood, and fecal samples of six groups of 10-12 day-old offspring were collected. The results demonstrated that maternal milk oligosaccharides and probiotics conferred protective effects against lipopolysaccharide (LPS)-induced immunosuppression in mice offspring by significantly enhancing the immune organ indexes, splenocyte proliferation, immunoglobulin (immunoglobulin G, A, M) production as well as improving the macrophage phagocytosis (p < .05). The abundance of Lactobacilli and Bifidobacteria in the feces of offspring mice in the intervention groups was significantly higher than that of the offspring mice in the control group (p < .05). These findings suggest that the combination of 2'-FL and Bifidobacterium Bb12 displayed synergistic interactions between the two components that could promote the development of the immune system of the offsprings and improve their microbiota through maternal ingestion.

White-light crosslinkable milk protein bioadhesive with ultrafast gelation for first-aid wound treatment.

BACKGROUND: Post-traumatic massive hemorrhage demands immediately available first-aid supplies with reduced operation time and good surgical compliance. In-situ crosslinking gels that are flexibly adapting to the wound shape have a promising potential, but it is still hard to achieve fast gelation, on-demand adhesion, and wide feasibility at the same time. METHODS: A white-light crosslinkable natural milk-derived casein hydrogel bioadhesive is presented for the first time. Benefiting from abundant tyrosine residues, casein hydrogel bioadhesive was synthesized by forming di-tyrosine bonds under white light with a ruthenium-based catalyst. We firstly optimized the concentration of proteins and initiators to achieve faster gelation and higher mechanical strength. Then, we examined the degradation, cytotoxicity, tissue adhesion, hemostasis, and wound healing ability of the casein hydrogels to study their potential to be used as bioadhesives. RESULT: Rapid gelation of casein hydrogel is initiated with an outdoor flashlight, a cellphone flashlight, or an endoscopy lamp, which facilitates its usage during first-aid and minimally invasive operations. The rapid gelation enables 3D printing of the casein hydrogel and excellent hemostasis even during liver hemorrhage due to section injury. The covalent binding between casein and tissue enables robust adhesion which can withstand more than 180 mmHg blood pressure. Moreover, the casein-based hydrogel can facilitate post-traumatic wound healing caused by trauma due to its biocompatibility. CONCLUSION: Casein-based bioadhesives developed in this study pave a way for broad and practical application in emergency wound management.

Scalable Milk-Derived Whey Protein Hydrogel as an Implantable Biomaterial.

There are limited naturally derived protein biomaterials for the available medical implants. High cost, low yield, and batch-to-batch inconsistency, as well as intrinsically differing bioactivity in some of the proteins, make them less beneficial as common implant materials compared to their synthetic counterparts. Here, we present a milk-derived whey protein isolate (WPI) as a new kind of natural protein-based biomaterial for medical implants. The WPI was methacrylated at 100 g bench scale, >95% conversion, and 90% yield to generate a photo-cross-linkable material. WPI-MA was further processed into injectable hydrogels, monodispersed microspheres, and patterned scaffolds with photo-cross-linking-based advanced processing methods including microfluidics and 3D printing. In vivo evaluation of the WPI-MA hydrogels showed promising biocompatibility and degradability. Intramyocardial implantation of injectable WPI-MA hydrogels in a model of myocardial infarction attenuated the pathological changes in the left ventricle. Our results indicate a possible therapeutic value of WPI-based biomaterials and give rise to a potential collaboration between the dairy industry and the production of medical therapeutics.

Real-Time In Situ Screening of Omega-7 Phospholipids in Marine Biological Resources Using an iKnife-Rapid-Evaporative-Ionization-Mass-Spectrometry-Based Lipidomics Phenotype.

Omega-7 (n-7) phospholipids were bioactive substances in marine animals. In this study, a fast lipidomics phenotyping approach for real-time in situ screening of n-7 phospholipids in five kinds of economic seafood, salmon, prawn, bluefin tuna, hairtail, and butterfish, was established using iKnife rapid evaporative ionization mass spectrometry (REIMS). The n-7 phospholipids were structurally characterized and quantitatively analyzed, and the profiles were statistically analyzed by multivariate recognition analysis. It indicated that the difference of n-7 phospholipids in seafood samples was significant (p < 0.05), with R2(cum) and Q2(cum) values of >0.9. The proportion of n-7 phospholipids in salmon was the highest (20.43%), followed by bluefin tuna, prawn, hairtail, and butterfish. The ions of m/z 742.54 (PC 16:1-18:1), 768.55 (PC 16:1-20:2), 697.48 (PE 16:1-18:1), and 699.48 (PE 16:1-18:0) were the main n-7 phospholipids. The effectiveness of iKnife REIMS was further verified by hydrophilic interaction chromatography mass spectrometry and gas chromatography. The results demonstrated that proposed iKnife REIMS was an excellent technique for front-line screening of n-7 phospholipids in a large variety of marine biological resources.

Lipidomics phenotyping of clam (Corbicula fluminea) through graphene/fibrous silica nanohybrids based solid-phase extraction and HILIC-MS analysis.

Polyunsaturated phospholipids are abundant in clam (Corbicula fluminea) but difficult to be fully extracted. Herein, graphene/fibrous silica (G/KCC-1) nanohybrids were synthesized, characterized, and applied for solid-phase extraction (SPE) of phospholipids in clam. The effectiveness of G/KCC-1 SPE was verified by hydrophilic interaction chromatography mass spectrometry (HILIC-MS) based lipidomics and statistical analysis. The ions of PE 16:0/18:1 (m/z 716.4), PC 16:0/20:5 (m/z 824.6) and etc. were regarded as the main difference among the crude lipids, acetone washed extract, and eluate of G/KCC-1 SPE. Finally, this method was validated in terms of linearity (R2 0.9965 to 0.9981), sensitivity (LOD 0.19-0.51 µg·mL-1 and LOQ 0.48 - 1.47 µg·mL-1), and precision (RSDintra-day ≤ 7.16% and RSDinter-day ≤ 7.30%). In conclusion, the G/KCC-1 SPE and HILIC-MS method was shown to be accurate and efficient in selective extracting and phenotyping phospholipids in C. fluminea.

Titania-coated fibrous silica (TiO2/KCC-1) core-shell microspheres based solid-phase extraction in clam (Corbicula fluminea) using hydrophilic interaction liquid chromatography and mass spectrometry.

Clam (Corbicula fluminea) contains significant amount of phospholipids, and the profile of phospholipids could be used as an index for nutrition evaluation. In this study, a titania-coated fibrous silica (TiO2/KCC-1) core-shell microsphere based solid-phase extraction (SPE) and hydrophilic interaction liquid chromatography mass spectrometry (HILIC-MS) method was developed to study the phospholipids in clam. The structure and morphology of TiO2/KCC-1 were characterized by transmission electron microscopy (TEM), scanning electron microscope (SEM), X-ray diffraction (XRD), and ultraviolet-visible diffuse reflectance spectroscopy (UV-vis DRS). The analytical results indicated that the phospholipid molecular species (PMS) could be extracted in a cleaner manner after TiO2/KCC-1 SPE, benefiting from the enhanced signals and decreased MS noise of phospholipids. Based on the principal component analysis (PCA) and partial least squares regression discriminant analysis (PLS-DA) models, the PMS with the most obvious beneficiaries of TiO2/KCC-1 SPE were identified, e.g. PC 16:0/20:5 (m/z 824.6), PC 16:0/22:6 (m/z 850.6), PE 18:1/18:2 (m/z 740.4), and PE 18:0/22:6 (m/z 790.5). Finally, the proposed method was validated to be sensitive and precise. This study provides an efficient tool to analyze the phospholipids in clam and the prospect to use this method for future applications in various samples.

The chemokine CXCL9 expression is associated with better prognosis for colorectal carcinoma patients.

The chemokine CXCL9 has been demonstrated to play an important role in the development of human malignancies. However, its prognostic significance in cancer patients remains unclear and less is known about its role in colonrectal carcinoma (CRC) patients. In this study, we found that the relative mRNA expression level of CXCL9 in primary colorectal tumor tissues was significantly higher than that in corresponding normal colon tissues. CXCL9 protein expression was also detected in 102 of 130 primary CRC patients by immunochemistry. Thus, CXCL9 might play a vital role in the progression of colorectal cancer. By analyzing the correlation between clinicopathological factors of patients and expression of CXCL9 protein, we showed that the expression of CXCL9 was significantly associated with tumor differentiation, tumor invasion, lymph node metastasis, distant metastasis, and vascular invasion, but not with other factors of CRC patients including age, gender, tumor location and tumor size. Furthermore, by performing Kaplan-Meier method as well as Cox's univariate and multivariate hazard regression model, we found that the higher the CXCL9 expression, the higher overall survival rate was observed, and CXCL9 expression was a significant independent prognostic factor for CRC patients. Therefore, CXCL9 is a useful predictor of better clinical outcome in CRC patients.