Assessing Leachable Cytotoxicity of 3D-Printed Polymers and Facile Detoxification Methods.

Additive manufacturing of polymers is gaining momentum in health care industries by providing rapid 3D printing of customizable designs. Yet, little is explored about the cytotoxicity of leachable toxins that the 3D printing process introduced into the final product. We studied three printable materials, which have various mechanical properties and are widely used in stereolithography 3D printing. We evaluated the cytotoxicity of these materials through exposing two fibroblast cell lines (human and mouse derived) to the 3D-printed parts, using overlay indirect contact assays. All the 3D-printed parts were measured toxic to the cells in a leachable manner, with flexible materials more toxic than rigid materials. Furthermore, we attempted to reduce the toxicity of the 3D-printed material by employing three treatment methods (further curing, passivation coating, and Soxhlet solvent extraction). The Soxhlet solvent extraction method was the most effective in removing the leachable toxins, resulting in the eradication of the material's toxicity. Passivation coating and further curing showed moderate and little detoxification, respectively. Additionally, mechanical testing of the materials treated with extraction methods revealed no significant impacts on its mechanical performances. As leachable toxins are broadly present in 3D-printed polymers, our cytotoxicity evaluation and reduction methods could aid in extending the selections of biocompatible materials and pave the way for the translational use of 3D printing.

A school-based public health model to reduce oral health disparities.

OBJECTIVES: Although dental decay is preventable, it remains the most common pediatric chronic disease. We describe a public health approach to implementing a scalable and sustainable school-based oral health program for low-income urban children. METHODS: The Los Angeles Trust for Children's Health, a nonprofit affiliated with the Los Angeles Unified School District, applied a public health model and developed a broad-based community-coalition to a) establish a District Oral Health Nurse position to coordinate oral health services, and b) implement a universal school-based oral health screening and fluoride varnishing program, with referral to a dental home. Key informant interviews and focus groups informed program development. Parent surveys assessed preventative oral health behaviors and access to oral health services. Results from screening exams, program costs and rates of reimbursement were recorded. RESULTS: From 2012 to 2015, six elementary schools and three dental provider groups participated. Four hundred ninety-one parents received oral health education and 89 served as community oral health volunteers; 3,399 screenings and fluoride applications were performed on 2,776 children. Sixty-six percent of children had active dental disease, 27 percent had visible tooth decay, and 6 percent required emergent care. Of the 623 students who participated for two consecutive years, 56 percent had fewer or no visible caries at follow-up, while only 17 percent had additional disease. Annual program cost was $69.57 per child. CONCLUSIONS: Using a broad based, oral health coalition, a school-based universal screening and fluoride varnishing program can improve the oral health of children with a high burden of untreated dental diseases.

Biofilms Benefiting Plants Exposed to ZnO and CuO Nanoparticles Studied with a Root-Mimetic Hollow Fiber Membrane.

Plants exist with a consortium of microbes that influence plant health, including responses to biotic and abiotic stress. While nanoparticle (NP)-plant interactions are increasingly studied, the effect of NPs on the plant microbiome is less researched. Here a root-mimetic hollow fiber membrane (HFM) is presented for generating biofilms of plant-associated microbes nurtured by artificial root exudates (AREs) to correlate exudate composition with biofilm formation and response to NPs. Two microbial isolates from field-grown wheat, a bacillus endophyte and a pseudomonad root surface colonizer, were examined on HFMs fed with AREs varying in N and C composition. Bacterial morphology and biofilm architecture were characterized using scanning electron microscopy (SEM) and atomic force microscopy (AFM) and responses to CuO and ZnO NP challenges of 300 mg/L evaluated. The bacillus isolate sparsely colonized the HFM. In contrast, the pseudomonad formed robust biofilms within 3 days. Dependent on nutrient sources, the biofilm cells produced extensive extracellular polymeric substances (EPS) and large intracellular granules. Pseudomonad biofilms were minimally affected by ZnO NPs. CuO NPs, when introduced before biofilm maturation, strongly reduced biofilm formation. The findings demonstrate the utility of the HFM root-mimetic to study rhizoexudate influence on biofilms of root-colonizing microbes but without active plant metabolism. The results will allow better understanding of how microbe-rhizoexudate-NP interactions affect microbial and plant health.

Cu from dissolution of CuO nanoparticles signals changes in root morphology.

Utilization of CuO nanoparticles (NPs) in agriculture, as fertilizers or pesticides, requires understanding of their impact on plant metabolism. Inhibition of root elongation by CuO NPs (>10 mg Cu/kg) occurred in wheat grown in sand. Morphological changes included root hair proliferation and shortening of the zones of division and elongation. The epidermal cells in the compressed root tip were abnormal in shape and file patterning but staining with SYTOX Blue did not reveal a general increase in epidermal cell death. Inhibition of root elongation and proliferation of root hair formation occurred also in response to exogenous indole acetic acid (IAA) supplied through tryptophan metabolism by the root-colonizing bacterium, Pseudomonas chlororaphis O6. Altered root morphology caused by the CuO NPs was likely due to release of Cu from dissolution at the root surface because similar changes occurred with Cu ions (≥6 mg/kg). Use of a fluorescent probe showed the accumulation of nitric oxide (NO), required for root hair formation, was not changed by the NPs. These findings suggested that dissolution of the NPs in the rhizosphere resulted levels of Cu that modified IAA distribution to causing root shortening but permitted NO cell signaling to promote root hair proliferation.

An indigoidine biosynthetic gene cluster from Streptomyces chromofuscus ATCC 49982 contains an unusual IndB homologue.

A putative indigoidine biosynthetic gene cluster was located in the genome of Streptomyces chromofuscus ATCC 49982. The silent 9.4-kb gene cluster consists of five open reading frames, named orf1, Sc-indC, Sc-indA, Sc-indB, and orf2, respectively. Sc-IndC was functionally characterized as an indigoidine synthase through heterologous expression of the enzyme in both Streptomyces coelicolor CH999 and Escherichia coli BAP1. The yield of indigoidine in E. coli BAP1 reached 2.78 g/l under the optimized conditions. The predicted protein product of Sc-indB is unusual and much larger than any other reported IndB-like protein. The N-terminal portion of this enzyme resembles IdgB and the C-terminal portion is a hypothetical protein. Sc-IndA and/or Sc-IndB were co-expressed with Sc-IndC in E. coli BAP1, which demonstrated the involvement of Sc-IndB, but not Sc-IndA, in the biosynthetic pathway of indigoidine. The yield of indigoidine was dramatically increased by 41.4 % (3.93 g/l) when Sc-IndB was co-expressed with Sc-IndC in E. coli BAP1. Indigoidine is more stable at low temperatures.

Metabolism of quercetin by Cunninghamella elegans ATCC 9245.

Incubation of quercetin with Cunninghamella elegans ATCC 9245 yielded three metabolites, including quercetin 3-O-ß-D-glucopyranoside, kaempferol 3-O-ß-D-glucopyranoside and isorhamnetin 3-O-ß-D-glucopyranoside. Glucosylation, O-methylation and dehydroxylation were involved in the process, among which dehydroxylation has never been found in Cunninghamella. Quercetin was completely metabolized in 72 h.

Generation of two new macrolactones through sequential biotransformation of dihydroresorcylide.

Biotransformation is an effective method to generate new derivatives from natural products. Combination of various enzymes or whole-cell biocatalysts creates new opportunities for natural product biosynthesis. Dihydroresorcylide (1) is a phytotoxic macrolactone from Acremonium aeae. It was first chlorinated at C-11 by an engineered Escherichia coli BL21-CodonPlus (DE3)-RIL/pJZ54 strain that overexpresses a fungal flavin-dependent halogenase, and subsequently glycosylated at 12-OH by Beauveria bassiana ATCC 7159, giving rise to a novel derivative, 11-chloro-4'-O-methyl-12-O-beta-D-glucosyl-dihydroresorcylide (3). Although 1 can be converted into a new 4'-O-methyl-glucosylated derivative 4 by B. bassiana, this product cannot be further chlorinated by E. coli BL21-CodonPlus (DE3)-RIL/pJZ54 to afford 3. The sequence of these two biotransformation steps was thus restricted and not interchangeable. This sequential biotransformation approach can be applied to other structurally similar natural products to create novel derivatives.

Specific inhibition of the halogenase for radicicol biosynthesis by bromide at the transcriptional level in Pochonia chlamydosporia.

Pochonia chlamydosporia produces radicicol (1), a potent antifungal and anticancer product. NaBr, but not NaF, NaCl or NaI, inhibited the biosynthesis of 1 in P. chlamydosporia in a dose-dependent manner, accompanied by the formation chlorine-lacking monocillins II-V (2-5), indicating that the dedicated halogenase, Rdc2 had been inhibited. RT-PCR analysis confirmed that transcription of rdc2 was selectively inhibited by Br(-), whereas the putative P450 epoxidase gene, rdc4, was not affected.