Ultrasmall Fe@Fe3O4 nanoparticles as T1-T2 dual-mode MRI contrast agents for targeted tumor imaging.

Targeted T1-T2 MRI contrast agents, which can eliminate the difficulty of image matching across multiple imaging instruments and permit specific localization of lesions, are promising candidates for more accurate diagnosis of tumors. In this study, ultrasmall Fe@Fe3O4 nanoparticles were designed and synthesized as T1-T2 dual-mode MRI contrast agents for accurate tumor imaging. First, to investigate the influence of nanoparticle size, Fe@Fe3O4 nanoparticles with diameters of 4, 8, and 12 nm were prepared, among which the 8 nm 3-(3,4-dihydroxyphenyl)propionic acid (DHCA)-modified nanoparticles exhibited the optimal T1-T2 dual-mode MRI performance. Next, to develop a tumor-targeted contrast agent, the DHCA-Fe@Fe3O4 nanoparticles were conjugated with the F56 peptide, which targets the vascular endothelial growth factor receptor, and the resulting F56-DHCA-Fe@Fe3O4 nanoparticles were found to exhibit good T1-T2 dual-mode imaging and tumor-targeting performance both in vitro and in vivo, indicating the nanoparticles represent a new research tool for accurate tumor diagnosis.

An up-converting phosphor technology-based lateral flow assay for point-of-collection detection of morphine and methamphetamine in saliva.

Morphine (Mop) and methamphetamine (Met) are highly addictive drugs worldwide. Point-of-collection testing (POCT) for drug-of-abuse screening is important in abuse/rehabilitation clinics and law-enforcement agencies. We established an up-converting phosphor technology-based lateral flow assay (UPT-LFA) as a point-of-collection testing (POCT) method, namely Mop-UPT-LFA and Met-UPT-LFA, for the detection of morphine and methamphetamine without complicated sample pre-treatment, respectively, in saliva. The sensitivities of the Mop-UPT-LFA and the Met-UPT-LFA were 5 and 10 ng mL-1 with accurate quantitation of 5-100 ng mL-1 and 10-250 ng mL-1 for morphine and methamphetamine, respectively, for a detection time of 15 min. In reference to the detection limits of 20 and 25 ng mL-1 for morphine and methamphetamine, respectively, in the Driving Under the Influence of Drugs, Alcohol and Medicines (DRUID) program of the European Union, the percentage test/control (T/C) ratio of the UPT-LFA between 2 and 15 min reached 101% and 86%, and the UPT-LFA produced accurate qualitative results in 2 min for 100 simulated-saliva samples with the exception of a few weakly positive samples. The sample and sample treating buffer were mixed and added to the test strip, and the test was conducted 15 min later. Although we found no significant difference between the UPT-LFA quantitative test and the liquid chromatography tandem mass spectrometry (LC-MS) test, compared with the latter, the UPT-LFA was substantially faster and had higher detection efficiency. The UPT-LFA showed more accurate qualitative results than the LC-MS for 50 simulated-saliva samples. The ease of operation, high sensitivity, and accuracy of the UPT-LFA make it a valid candidate POCT method for drug-of-abuse screening.

Fe3O4 assembly for tumor accurate diagnosis by endogenous GSH responsive T2/T1 magnetic relaxation conversion.

Superparamagnetic iron oxide nanoparticles with high magnetization strength and good biological safety have been widely used as magnetic resonance imaging (MRI) contrast agents for tumors. However, the accuracy of tumor diagnosis is still low due to the lack of tumor targeting and the interference signals from normal tissues. Endogenous substances in tumor (such as high levels of GSH and pH) stimuli-responsive contrast agents could offer higher sensitivity for tumor diagnosis. Herein, based on the characteristic of overexpression of GSH in tumors, we propose an ultra-small Fe3O4 assembly as an endogenous GSH responsive MRI contrast agent. The ultra-small superparamagnetic Fe3O4 are bonded to the crosslinker cystamine to synthesize Fe3O4 nanoclusters, which exhibit a T2 imaging effect. When the contrast agent reaches the tumor tissue, the disulfide bond in cystamine is induced by GSH to break, the Fe3O4 nanoclusters are disassembled into ultra-small Fe3O4 nanoparticles, and the relaxation signal changes from T2 to T1, which is helpful for accurate diagnosis of tumors. In vivo experiments have shown that Fe3O4 nanoclusters can rapidly respond to overexpressed GSH in tumor sites for T2/T1 switchable imaging. This work not only designed an endogenous GSH responsive platform through simple synthesis methods, but also improved the accuracy of tumor diagnosis through the transformation of T2/T1 MRI signals.

Late-onset Hearing Loss From Congenital Cytomegalovirus Infection After Newborn Period in a Highly Immune Population in China.

After following 141 children with likely asymptomatic congenital cytomegalovirus infection in a highly immune population in China, four children (2.8%) were found to have late-onset hearing loss. No maternal or childhood factors, except higher saliva cytomegalovirus viral load at birth (P = 0.03), were associated with increased risk of developing a hearing loss.

GGTA1/iGb3S Double Knockout Mice: Immunological Properties and Immunogenicity Response to Xenogeneic Bone Matrix.

BACKGROUND: Animal tissues and tissue-derived biomaterials are widely used in the field of xenotransplantation and regenerative medicine. A potential immunogenic risk that affects the safety and effectiveness of xenografts is the presence of remnant α-Gal antigen (synthesized by GGTA1 or/and iGb3S). GGTA1 knockout mice have been developed as a suitable model for the analysis of anti-Gal antibody-mediated immunogenicity. However, we are yet to establish whether GGTA1/iGb3S double knockout (G/i DKO) mice are sensitive to Gal antigen-positive xenoimplants. METHODS: α-Gal antigen expression in the main organs of G/i DKO mice or bovine bone substitutes was detected via a standardized ELISA inhibition assay. Serum anti-α-Gal antibody titers of G/i DKO mice after immunization with rabbit red blood cells (RRBC) and implantation of raw lyophilized bone substitutes (Gal antigen content was 8.14 ± 3.17 × 1012/mg) or Guanhao Biotech bone substitutes (50% decrease in Gal antigen relative to the raw material) were assessed. The evaluation of total serum antibody, inflammatory cytokine, and splenic lymphocyte subtype populations and the histological analysis of implants and thymus were performed to systematically assess the immune response caused by bovine bone substitutes and bone substitute grafts in G/i DKO mice. RESULTS: α-Gal epitope expression was reduced by 100% in the main organs of G/i DKO mice, compared with their wild-type counterparts. Following immunization with RRBC, serum anti-Gal antibody titers of G/i DKO mice increased from 80- to 180-fold. After subcutaneous implantation of raw lyophilized bone substitutes and Guanhao Biotech bone substitutes into G/i DKO mice, specific anti-α-Gal IgG, anti-α-Gal IgM, and related inflammatory factors (IFN-γ and IL-6) were significantly increased in the raw lyophilized bone substitute group but showed limited changes in the Guanhao Biotech bone substitute group, compared with the control. CONCLUSION: G/i DKO mice are sensitive to Gal antigen-positive xenogeneic grafts and can be effectively utilized for evaluating the α-Gal-mediated immunogenic risk of xenogeneic grafts.

Xenogeneic bone matrix immune risk assessment using GGTA1 knockout mice.

Homeotransplantation of bones for replacement therapy have been demonstrated reliably in clinical data. However, human donor bones applicable for homeotransplantation are in short supply, which facilitates the search for suitable alternatives, such as xenografts grafts. The α-Gal antigen-related immune risk of xenografts directly affects the safety and effectiveness of the biomaterials and limits their applications in the clinic. The immune risk can be prevented by depletion or breaking anti-Gal antibody prior to transplant. Therefore, how to assess the immune risk of the bone substitutes and select the reliable animal research model become extremely important. In this study, we prepared lyophilized bone substitutes (T1) and Guanghao Biotech bone substitutes (T2, animal-derived biomaterials with α-Gal antigen decreased), aimed to assess the immune risk of xenografts bone substitutes on GGTA1 knockout mice. The α-Gal antigen contents of T1 and T2 were firstly detected by ELISA method in vitro. The bone substitutes were then implanted subcutaneously into GGTA1 knockout mice for 2, 4 and 12 weeks, respectively. The total serum antibody levels, anti-α-Gal antibody levels, inflammatory cytokine and splenic lymphocyte surface molecules were detected and histology analysis of skin and thymus were performed to systematically evaluate the immune response caused by the T1 and T2 bone substitutes in mice. In vitro results showed that the amount of α-Gal epitopes in T1 bone substitutes was significantly higher than T2 bone substitutes, and the clearance rate of α-Gal antigen in T2 bone substitutes achieved about 55.6%. Results of antibody level in vivo showed that the T1 bone substitutes group possessed significantly higher total IgG, IgM, IgA and anti-α-Gal IgG levels than T2 and control group, while T2 group showed no significant changes of these indexes compared with control. In terms of inflammatory cytokines, T1 bone substitutes showed evidently higher levels of IL-4, IL-12P70 and IL-10 than T2 and control, while T2 group was comparable to control. No changes in the levels of splenic lymphocyte surface molecules were found in the three groups (T1, T2 and control group) during the experimental periods. The pathological results demonstrated that the inflammatory response in T2 group was lighter than the T1 group, which was in accordance with the inflammatory cytokines levels. The above results indicated that the process of antigen removal effectively reduced the α-Gal antigens content in T2 bone substitutes, which caused little immune response in vivo and could be used as bone healing materials. This study also demonstrated that GGTA1 knockout mice can be used as a routine tool to assess the immune risk of animal-derived biomaterials.

Viral Loads in Congenital Cytomegalovirus Infection From a Highly Immune Population.

Among newborns with congenital cytomegalovirus (CMV) infection from China, there was no difference in CMV viral load in saliva specimens dried and stored at room temperature compared with those kept wet and stored cold, even after longer storage time for the former than the later (74 vs 58 days, P = .02).

Cohort study on maternal cytomegalovirus seroprevalence and prevalence and clinical manifestations of congenital infection in China.

Congenital cytomegalovirus (CMV) infection is the leading viral cause of birth defects and developmental disabilities in developed countries. However, CMV seroprevalence and burden of congenital CMV infection are not well defined in China.Cohort of newborns from 5 birthing hospitals in 2 counties of Shandong Province, China, were enrolled from March 2011 to August 2013. Dried blood spots (DBS) and saliva were collected within 4 days after birth for IgG testing for maternal seroprevalence and real-time PCR testing for congenital CMV infection, respectively.Among 5020 newborns tested for CMV IgG, 4827 were seropositive, resulting in CMV maternal seroprevalence of 96.2% (95% confidence interval [CI]:95.6%-96.7%). Of the 10,933 newborns screened for congenital CMV infection, 75 had CMV detected, resulting in an overall prevalence of 0.7% (95% CI: 0.5%-0.9%), with prevalences of 0.4% (14/3995), 0.6% (66/10,857), and 0.7% (52/7761) for DBS, wet saliva, and dried saliva specimens screened, respectively. Prevalence of congenital CMV infection decreased with increasing maternal age (0.9%, 0.6%, and 0.3% among newborns delivered from mothers aged 16-25, 26-35, and >35 years, respectively; P = 0.03), and was higher among preterm infants than full term infants (1.3% vs 0.6%, P = 0.04), infants with intrauterine growth restriction (IUGR) than those without (1.8% vs 0.7%, P = 0.03), and twins or triplets than singleton pregnancies (2.8% vs 0.7%, P = 0.04). None of the 75 newborns exhibited symptomatic congenital CMV infection, and there was no difference in clinical characteristics and newborn hearing screening results between infants with and without congenital CMV infection at birth.Congenital CMV infection prevalence was lower and the clinical manifestations were milder in this relatively developed region of China compared to populations from other countries with similarly high maternal seroprevalence. Follow-up on children with congenital CMV infection will clarify the burden of disabilities from congenital CMV infection in China.

Particle-enhanced turbidimetric immunoassay for determination of serum neutrophil gelatinase-associated lipocalin on the Roche Cobas c501 analyzer.

OBJECTIVE: Neutrophil gelatinase-associated lipocalin (NGAL) has been reported to be a good marker for tubular damage and acute kidney injury. The aim of this study was to develop a high throughput assay for the quantification of serum NGAL (sNGAL). METHODS: Imprecision, interference, linearity, recovery, and reference values were evaluated on Cobas c501. RESULTS: The assay was linear over the dynamic range of the study (R(2)=0.9988). The total assay imprecision was below 5%. The assay recovery was estimated at 98.89%-102.61%. The assay displayed a good linearity over the range from 35 µg/L to 4250 µg/L. A typical high-dose hook effect was observed for the assay at NGAL concentration>28,800 µg/L. No interference was observed with hemoglobin ≤ 5 g/L, bilirubin ≤ 0.3g/L, vitamin C ≤ 0.5 g/L, sodium heparin≤ 5 g/L and intralipid ≤ 1%. The 95th centile for serum NGAL was <122.57 µg/L from 454 healthy donors. There were no gender-related differences for serum NGAL. There were significant age-related differences between the 21-44 and 45-75 year categories for serum NGAL. The reference value for sNGAL was <116.52 µg/L in the 21-44 year group and <126.9 µg/L in the 45-75 year group. CONCLUSIONS: The NGAL assay verified to be a reliable assay with convenient performance characteristics. The assay improves and simplifies the laboratory workload.