Effects of three products from Antarctic krill on the nitrogen balance, growth, and antioxidation status of rats.

A few studies conducted over the past few decades have demonstrated the health benefits of a diet rich in marine products, but limited studies have investigated the effects of different krill products on the nitrogen balance and their potential health benefits. In our study, after a 14-day acclimation period, 50 female Sprague-Dawley rats were randomly assigned to five groups, each of which was fed a different diet, for 28 days. We then evaluated the effect of krill protein complex (KPC), krill powder, and defatted krill powder on the nitrogen balance, growth, and antioxidant activity through analyses of MDA, CAT, GSH-Px, and T-SOD. An in vivo analysis suggested that the nitrogen retention rate, protein digestibility, and bioutilization of krill products were equal to those of casein. Moreover, the KPC diet resulted in the highest nitrogen intake and retention among the groups, and the biological value and net protein utilization obtained with KPC were higher than those obtained with defatted krill powder, which was consistent with the weight gains observed for these two groups. The hematological test also showed that KPC contributed to the production of functional proteins in the body. The antioxidant activity analysis indicated that higher GSH-Px and T-SOD activities were obtained with krill products and KPC, respectively, compared with casein. The results from this study suggested that krill proteins could promote growth and improve the antioxidant status of an organism. Although further studies on the safety of krill products for human consumption are needed, this work provides insights into the use of krill proteins as a potential substitute for other proteins and restructured foods.

Evaluating the efficacy of a ferrous-ion-chelating peptide from Alaska pollock frame for the improvement of iron nutritional status in rats.

This study aimed to investigate the effects of ferrous-ion-chelating peptides from Alaska pollock frames (APFP-Fe) on iron deficiency in anaemic rats. We hydrolysed the Alaska pollock frames to obtain a peptide with an average molecular weight of 822 Da. The bioavailability of APFP-Fe was tested using animal experiments. Wistar rats were randomly divided into six groups: an iron deficiency control group, a normal control group, and iron deficiency groups treated with ferrous sulfate (FeSO4) or low-, medium-, or high-dose APFP-Fe. Rats in the iron deficiency groups were fed an iron-deficient diet to establish the iron deficiency anaemia (IDA) model. After the model was established, different iron supplements were given to rats once per day via intragastric administration for 21 days. The results showed that APFP-Fe had restorative effects, returning the body weight, weight gain, height, and haematological parameters in IDA rats to normal levels. In addition, compared with FeSO4, APFP-Fe promoted significant weight gain and effectively improved haemoglobin, serum iron and transferrin levels, and recovery of the capacity of iron binding with transferrin, especially at the medium and high doses. These findings suggest that APFP-Fe is an effective source of iron for improving the iron nutritional status in IDA rats and shows promise as a new source of iron supplementation.

Regeneration of different types of tissues depends on the interplay of stem cells-laden constructs and microenvironments in vivo.

The ability of repair and regeneration of tissues or organs has been significantly improved by using biomaterials-based constructs. Our previous studies found the regeneration of both articular cartilage and subchondral bone by implantation of a poly(lactide-co-glycolide) (PLGA)/fibrin gel/bone marrow stem cells (BMSCs)/(lipofectamine/pDNA-transforming growth factor (TGF)-ß1) construct in vivo, without the step of pre-induced differentiation of the laden stem cells in vitro. To substantiate the ability to regenerate multi-types of tissues by the same constructs, in this study the constructs were implanted into three types of tissues or tissue defects in vivo, including subcutaneous fascia layer, and ear cartilage and eyelid tarsal plate defects. The ear cartilage and eyelid tarsal plate defects were fully regenerated 8 w post-implantation, showing a similar morphology to the corresponding native tissues. In the neo ear cartilage, abundant chondrocytes with obvious lacunas and cartilage-specific extracellular matrices (ECMs) were found. Neo eyelid tarsal plate with mature meibomian gland acinar units was regenerated. Furthermore, expressions of the ECMs-specific genes and proteins, as well as the cell behavior modulatory factors, Sry related HMG box 9 (Sox9) and TGF-ß1 were significantly up-regulated in the regenerated ear cartilages and eyelid tarsal plate than those in the subcutaneously implanted constructs, which were filled with fibrocytes, inflammatory cells, obvious vascularization and slight ECMs deposition. These results confirm firmly the ability to regenerate multi-types of tissues by a stem cells-laden construct via adapting to the microenvironments of corresponding tissues.

Regeneration of the Osteochondral Defect by a Wollastonite and Macroporous Fibrin Biphasic Scaffold.

Osteochondral defect refers to the damage of cartilage as well as subchondral bone. Cartilage tissue engineering focusing on the regeneration of cartilage and disregarding the subchondral bone always leads to partial regeneration of the damage, resulting in poor mechanical and physiological properties. A scaffold suitable for in situ inductive regeneration of both types of tissues is urgently needed. In this study, a biphasic scaffold integrated by macro-porous fibrin and 3D-printed wollastonite (containing 8% MgSiO3 (CS-Mg8)) scaffolds, either preloaded with rabbit bone marrow mesenchymal stem cells (BMSCs) or not, were fabricated and used to repair osteochondral defects in vivo (full thickness osteochondral defects in rabbits, 4 mm in diameter and 4 mm in depth with bone marrow blood effusion). The fibrin scaffold had a pore size of 100-200 µm, and was degraded gradually and reached weight loss over 80% at 28 days. The presence of BMSCs could accelerate the degradation rate. BMSCs could well proliferate in the fibrin scaffold along with time prolongation. The CS-Mg8 scaffold possessed a regular structure of cross stacked CS-Mg8 rods, and was degraded rather slowly with a mass loss of 8.5% at 28 days. BMSCs adhered and showed well spreading on the CS-Mg8 scaffold, without apparent proliferation in vitro. In vivo transplantation of the biphasic scaffolds, either preloaded with BMSCs or not, could induce the regeneration of both cartilage and subchondral bone to a great extent. Loading of BMSCs enabled better regeneration of cartilage layer, leading to smoother macroscopic appearance, good integrity with surrounding tissue and tide mark formation. However, no significant difference in bone formation and in gene expression was found with and without BMSCs loading.

Macrophages of Different Phenotypes Influence the Migration of BMSCs in PLGA Scaffolds with Different Pore Size.

Immune modulation of the scaffolds not only reduces the host immunological rejection response, but also improves the regenerative cell migration into the scaffolds. Herein a convenient immune modulation of poly(lactide-co-glycolide) (PLGA) scaffold is applied with macrophages of different phenotypes to evaluate its influence on the migration behavior of bone marrow mesenchymal stem cells (BMSCs). With pro-inflammatory macrophages (M1) pre-loading, BMSCs migrate significantly faster into the PLGA scaffold, compared with those in the control scaffold or pre-seeded with inactivated macrophages (M0). The pore size of PLGA scaffolds is found to take a more important role, as the one with a larger pore size significantly enhance migration of BMSCs no matter the pre-seeding of macrophages. The enhanced cell migration in the macrophage-modulated scaffold can provide a new protocol for in situ tissue regeneration by recruiting endogenous cells.

Regeneration of osteochondral defects in vivo by a cell-free cylindrical poly(lactide-co-glycolide) scaffold with a radially oriented microstructure.

A scaffold with an oriented porous architecture to facilitate cell infiltration and bioactive interflow between neo-host tissues is of great importance for in situ inductive osteochondral regeneration. In this study, a poly(lactide-co-glycolide) (PLGA) scaffold with oriented pores in its radial direction was fabricated via unidirectional cooling of the PLGA solution in the radial direction, following with lyophilization. Micro-computed tomography evaluation and scanning electron microscopy observation confirmed the radially oriented microtubular pores in the scaffold. The scaffold had porosity larger than 90% and a compressive modulus of 4 MPa in a dry state. Culture of bone marrow stem cells in vitro revealed faster migration and regular distribution of cells in the poly(lactide-co-glycolide) scaffold with oriented pores compared with the random PLGA scaffold. The cell-free oriented macroporous PLGA scaffold was implanted into rabbit articular osteochondral defect in vivo for 12 weeks to evaluate its inductive tissue regeneration function. Histological analysis confirmed obvious tide mark formation and abundant chondrocytes distributed regularly with obvious lacunae in the cartilage layer. Safranin O-fast green staining showed an obvious boundary between the two layers with distinct staining results, indicating the simultaneous regeneration of the cartilage and subchondral bone layers, which is not the case for the random poly(lactide-co-glycolide) scaffold after the same implantation in vivo. The oriented macroporous PLGA scaffold is a promising material for the in situ inductive osteochondral regeneration without the necessity of preseeding cells.

Induced migration of endothelial cells into 3D scaffolds by chemoattractants secreted by pro-inflammatory macrophages in situ.

Cell migration in scaffolds plays a crucial role in tissue regeneration, which can better mimic cell behaviors in vivo. In this study, a novel model has been proposed on controlling 3D cell migration in porous collagen-chitosan scaffolds with various pore structures under the stimulation of inflammatory cells to mimic the angiogenesis process. Endothelial cells (ECs) cultured atop the scaffolds in the Transwell molds which were placed into a well of a 24-well culture plate were promoted to migrate into the scaffolds by chemoattractants such as vascular endothelial growth factor (VEGF) and tumor necrosis factor-alpha (TNF-α) secreted by the pro-inflammatory macrophages incubated in the well culture plate. The phenotype of macrophages was mediated by 50 ng/ml interferon-gamma (IFN-γ) and different concentrations of lipopolysaccharide (LPS, 150-300 ng/ml). The cell migration depth had a positive correlation with LPS concentration, and thereby the TNF-α concentration. The ECs migrated easier to a deeper zone of the scaffolds prepared at - 10ºC (187 µm in pore diameter) than that at - 20ºC (108 µm in pore diameter) as well. The method provides a useful strategy to study the 3D cell migration, and is helpful to reveal the vascularization process during wound healing in the long run.

Cell-Free HA-MA/PLGA Scaffolds with Radially Oriented Pores for In Situ Inductive Regeneration of Full Thickness Cartilage Defects.

A bioactive scaffold with desired microstructure is of great importance to induce infiltration of somatic and stem cells, and thereby to achieve the in situ inductive tissue regeneration. In this study, a scaffold with oriented pores in the radial direction is prepared by using methacrylated hyaluronic acid (HA-MA) via controlled directional cooling of a HA-MA solution, and followed with photo-crosslinking to stabilize the structure. Poly(lactide-co-glycolide) (PLGA) is further infiltrated to enhance the mechanical strength, resulting in a compressive modulus of 120 kPa. In vitro culture of bone marrow stem cells (BMSCs) reveals spontaneous cell aggregation inside this type of scaffold with a spherical morphology. In vivo transplantation of the cell-free scaffold in rabbit knees for 12 w regenerates simultaneously both cartilage and subchondral bone with a Wakitani score of 2.8. Moreover, the expression of inflammatory factor interleukin-1ß (IL-1ß) is down regulated, although tumor necrosis factor-α (TNF-α) is remarkably up regulated. With the anti-inflammatory, bioactive properties and good restoration of full thickness cartilage defect in vivo, the oriented macroporous HA-MA/PLGA hybrid scaffold has a great potential for the practical application in the in situ cartilage regeneration.

Cell-free macro-porous fibrin scaffolds for in situ inductive regeneration of full-thickness cartilage defects.

Macro-porous fibrin scaffolds with regular and adjustable inter-connective pores were fabricated through a porogen-leaching method for the in situ inductive regeneration of full-thickness cartilage defects in vivo. In vitro tests proved the survival and proliferation of BMSCs in the scaffolds. In vivo repair experiment was conducted by implantation of the cell-free macro-porous fibrin scaffolds into full-thickness cartilage defects (4 mm in diameter and 4 mm in depth with bone marrow blood effusion) of New Zealand white rabbits for 6 and 12 w. The neo-cartilage integrated well with the surrounding cartilage as well as the subchondral bone. Immunochemical and GAG staining revealed the abundant deposition of type II collagen and GAGs in the neo-cartilage after regeneration for 12 w. Histological score of the regenerated tissues was 2.6. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and western blotting (WB) revealed that the cartilage-related genes and proteins were significantly up-regulated compared with those of the normal cartilage. With the cell-free advantage and positive restoration of full-thickness cartilage defect in vivo, the fibrin scaffold is shelf-ready and is expected to be conveniently used in clinics.