Improving In Vitro Culture of Human Male Fetal Germ Cells

Male human fetal germ cells (hFGCs) give rise to spermatogonial stem cells (SSCs), which are the adult precursors of the male gametes. Human SSCs are a promising (autologous) source of cells for male fertility preservation; however, in contrast to mouse SSCs, we are still unable to culture them in the long term. Here, we investigated the effect of two different culture media and four substrates (laminin, gelatin, vitronectin and matrigel) in the culture of dissociated second trimester testes, enriched for hFGCs. After 6 days in culture, we quantified the presence of POU5F1 and DDX4 expressing hFGCs. We observed a pronounced difference in hFGC number in different substrates. The combination of gelatin-coated substrate and medium containing GDNF, LIF, FGF2 and EGF resulted in the highest percentage of hFGCs (10% of the total gonadal cells) after 6 days of culture. However, the vitronectin-coated substrate resulted in a comparable percentage of hFGCs regardless of the media used (3.3% of total cells in Zhou-medium and 4.8% of total cells in Shinohara-medium). We provide evidence that not only the choices of culture medium but also choices of the adequate substrate are crucial for optimizing culture protocols for male hFGCs. Optimizing culture conditions in order to improve the expansion of hFGCs will benefit the development of gametogenesis assays in vitro.

Investigation into Effect of Natural Shellac on the Bonding Strength of Magnesium Substituted Hydroxyapatite Coatings Developed on Ti6Al4V Substrates

The bioactive and biocompatible properties of hydroxyapatite (HA) promote the osseointegration process. To enhance other bio-functions of HA such as improving the antibacterial property of the implant, increasing the rate of cell proliferation, or improving tissue generation capability, HA is substituted with many elements such as Zn, Cl, Ba, Fe, Cu, Ag, Sr, F, Na, etc. This study reports development of Magnesium substituted HA (Mg-HA) coatings on Ti6Al4V substrates using the dip coating technique. To improve the adhesion and stability of the coating, an intermediate layer of shellac was applied between the coating and Ti6Al4V substrate. The dip coating process parameters were optimized using the Taguchi technique and it was found that dipping time of 35 s and 13% w/w of shellac concentration provided the maximum bonding strength of 12.5 MPa. The biocompatibility, dissolution, and corrosion study of the developed coating using the optimal parameters obtained were carried out in this study. An improvement in cell growth and cell proliferation was observed in the extract medium prepared from coated substrates. Release of Ca, P and Mg ions from the surface of the coated substrate into the simulated body fluid (SBF) was found to be almost constant which shows the stability of the thin film coating formed. The Mg-HA coated substrate also exhibited better corrosion resistance than the uncoated substrate.

Polydopamine-based biofunctional substrate coating promotes mesenchymal stem cell migration

Rapid migration of mesenchymal stem cells (MSCs) on device surfaces could support in vivo tissue integration and might facilitate in vitro organoid formation. Here, polydopamine (PDA) is explored as a biofunctional coating to effectively promote MSC motility. It is hypothesized that PDA stimulates fibronectin deposition and in this way enhances integrin-mediated migration capability. The random and directional cell migration was investigated by time-lapse microscopy and gap closure assay respectively, and analysed with softwares as computational tools. A higher amount of deposited fibronectin was observed on PDA substrate, compared to the non-coated substrate. The integrin β1 activation and focal adhesion kinase (FAK) phosphorylation at Y397 were enhanced on PDA substrate, but the F-actin cytoskeleton was not altered, suggesting MSC migration on PDA was regulated by integrin initiated FAK signalling. This study strengthens the biofunctionality of PDA coating for regulating stem cells and offering a way of facilitating tissue integration of devices. Graphic abstract Polydopamine-coated substrate induces increased fibronectin deposition of mesenchymal stem cells, and promotes cell migration via integrin-initiated FAK signaling, compared to non-coated polystyrene-based standard tissue culture surface. In this way, multifunctional PDA coating could support in vivo tissue integration on implant surface and promote in vitro organoid formation.

Drug-Releasing Antibacterial Coating Made from Nano-Hydroxyapatite Using the Sonocoating Method

Medical implant use is associated with a risk of infection caused by bacteria on their surface. Implants with a surface that has both bone growth-promoting properties and antibacterial properties are of interest in orthopedics. In the current study, we fabricated a bioactive coating of hydroxyapatite nanoparticles on polyether ether ketone (PEEK) using the sonocoating method. The sonocoating method creates a layer by immersing the object in a suspension of nanoparticles in water and applying a high-power ultrasound. We show that the simple layer fabrication method results in a well-adhering layer with a thickness of 219 nm to 764 nm. Dropping cefuroxime sodium salt (Cef) antibiotic on the coated substrate creates a layer with a drug release effect and antibacterial activity against Staphylococcus aureus. We achieved a concentration of up to 1 mg of drug per cm2 of the coated substrate. In drug release tests, an initial burst was observed within 24 h, accompanied by a linear stable release effect. The drug-loaded implants exhibited sufficient activity against S. aureus for 24 and 168 h. Thus, the simple method we present here produces a biocompatible coating that can be soaked with antibiotics for antibacterial properties and can be used for a range of medical implants.

Antibacterial coating of titanium-doped hydroxyapatite nanoparticles on a polymer substrate

Calcined and dispersible titanium-doped hydroxyapatite (Ti-HAp) nanoparticles at different [Ti/(Ca+Ti)] atomic ratios (0.3, 0.4, and 0.5) were prepared using an anti-sintering method. The Ti substitution ratios of the HAp structures in the feed of Ti-HAp preparation were approximately 80%. Ti-HAp nanoparticles were coated on polyethylene terephthalate (PET) sheets through polyacrylic acid graft-polymers. The PET substrate was almost completely covered with monolayer nanoparticles (over 95%). Antibacterial activity of coated Ti-HAp was calculated from the survival ratio of the bacteria, Staphylococcus aureus, after ultraviolet (UV) irradiation at 312 nm and 6.4 mW/cm2 for 30 s. The number of S. aureus on the Ti-HAp coated substrate decreased by 43% compared to those on the original PET and normal HAp coatings as negative controls. The antibacterial activity of Ti-HAp coated substrate was, furthermore, no statistically difference with TiO2 sheet as a positive control.

Hotspot generation for unique identification with nanomaterials

Nanoscale variations in the structure and composition of an object are an enticing basis for verifying its identity, due to the physical complexity of attempting to reproduce such a system. The biggest practical challenge for nanoscale authentication lies in producing a system that can be assessed with a facile measurement. Here, a system is presented in which InP/ZnS quantum dots (QDs) are randomly distributed on a surface of an aluminium-coated substrate with gold nanoparticles (Au NPs). Variations in the local arrangement of the QDs and NPs is shown to lead to interactions between them, which can suppress or enhance fluorescence from the QDs. This position-dependent interaction can be mapped, allowing intensity, emission dynamics, and/or wavelength variations to be used to uniquely identify a specific sample at the nanoscale with a far-field optical measurement. This demonstration could pave the way to producing robust anti-counterfeiting devices.

Solid-phase colorimetric sensor for hypochlorite

By taking advantage of the powerful oxidation property of hypochlorite (OCl−), we developed a solid-phase colorimetric sensor for the detection of OCl− based on 13 nm AuNPs immobilized on a 3-aminopropyltriethoxysilane APTES-coated substrate.

AuNP array coated substrate for sensitive and homogeneous SERS-immunoassay detection of human immunoglobulin G

An AuNP array coated substrate was developed for the SERS-immunoassay detection of human immunoglobulin G.