Mice lacking C1q or C3 show accelerated rejection of minor H disparate skin grafts and resistance to induction of tolerance

Complement activation is known to have deleterious effects on organ transplantation. On the other hand, the complement system is also known to have an important role in regulating immune responses. The balance between these two opposing effects is critical in the context of transplantation. Here, we report that female mice deficient in C1q (C1qa(−/−)) or C3 (C3(−/−)) reject male syngeneic grafts (HY incompatible) at an accelerated rate compared with WT mice. Intranasal HY peptide administration, which induces tolerance to syngeneic male grafts in WT mice, fails to induce tolerance in C1qa(−/−) or C3(−/−) mice. The rejection of the male grafts correlated with the presence of HY D(b)Uty-specific CD8(+) T cells. Consistent with this, peptide-treated C1qa(−/−) and C3(−/−) female mice rejecting male grafts exhibited more antigen-specific CD8(+)IFN-γ(+) and CD8(+)IL-10(+) cells compared with WT females. This suggests that accumulation of IFN-γ- and IL-10-producing T cells may play a key role in mediating the ongoing inflammatory process and graft rejection. Interestingly, within the tolerized male skin grafts of peptide-treated WT mice, IFN-γ, C1q and C3 mRNA levels were higher compared to control female grafts. These results suggest that C1q and C3 facilitate the induction of intranasal tolerance

Clinical presentation, auscultation recordings, ultrasonographic findings and treatment response of 12 adult cattle with chronic suppurative pneumonia: case study

Auscultation is considered the critical component of the veterinary clinical examination for the diagnosis of bovine respiratory disease but the accuracy with which adventitious sounds reflect underlying lung pathology remains largely unproven. Modern portable ultrasound machines provide the veterinary practitioner with an inexpensive, non-invasive tool with which to examine the pleural surfaces and superficial lung parenchyma. Simultaneous recording of sounds overlying normal lung and defined pathology allows critical assessment of auscultated sounds in the same animal removing confounding factors such as respiratory rate and thickness of the chest wall (body condition). Twelve cows, referred to the University of Edinburgh Veterinary School, were diagnosed with chronic suppurative pneumonia and enrolled into this prospective study to record and monitor lung sounds, ultrasonographic findings, and response to a standardised antibiotic treatment regimen. Most cows (8/12) had a normal rectal temperature on presentation but all cows had received antibiotic therapy at some time in the previous two weeks and six animals were receiving antibiotic treatment upon admission. All cattle were tachypnoeic (>40 breaths per minute) with frequent and productive coughing, halitosis, and a purulent nasal discharge most noticeable when the head was lowered. Ultrasonographic examination of the chest readily identified pathological changes consistent with severe lung pathology subsequently confirmed as chronic suppurative pneumonia in four cows at necropsy; eight cows recovered well after antibiotic treatment and were discharged two to six weeks after admission. It proved difficult to differentiate increased audibility of normal lung sounds due to tachypnoea from wheezes; coarse crackles were not commonly heard. In general, sounds were reduced in volume over consolidated lung relative to normal lung tissue situated dorsally. Rumen contraction sounds were commonly transmitted over areas of lung pathology. Trueperella (formerly Arcanobacterium) pyogenes was isolated from three of four lung tissue samples at necrospy. Treatment with procaine penicillin for 42 consecutive days resulted in marked improvement with return to normal appetite and improvement in body condition in 8 of 12 cows (67%) where lesions did not extend more than 10-15 cm above the level of the olecranon on both sides of the chest

Phenotypic Spectrum of Seizure Disorders in MBD5-Associated Neurodevelopmental Disorder

OBJECTIVE: To describe the phenotypic spectrum in patients with MBD5-associated neurodevelopmental disorder (MAND) and seizures; features of MAND include intellectual disability, epilepsy, psychiatric features of aggression and hyperactivity, and dysmorphic features including short stature and microcephaly, sleep disturbance, and ataxia. METHODS: We performed phenotyping on patients with MBD5 deletions, duplications, or point mutations and a history of seizures. RESULTS: Twenty-three patients with MAND and seizures were included. Median seizure onset age was 2.9 years (range 3 days–13 years). The most common seizure type was generalized tonic-clonic; focal, atypical absence, tonic, drop attacks, and myoclonic seizures occurred frequently. Seven children had convulsive status epilepticus and 3 nonconvulsive status epilepticus. Fever, viral illnesses, and hot weather provoked seizures. EEG studies in 17/21 patients were abnormal, typically showing slow generalized spike-wave and background slowing. Nine had drug-resistant epilepsy, although 3 eventually became seizure-free. All but one had moderate-to-severe developmental impairment. Epilepsy syndromes included Lennox-Gastaut syndrome, myoclonic-atonic epilepsy, and infantile spasms syndrome. Behavioral problems in 20/23 included aggression, self-injurious behavior, and sleep disturbance. CONCLUSION: MBD5 disruption may be associated with severe early childhood-onset developmental and epileptic encephalopathy. Because neuropsychiatric dysfunction is common and severe, it should be an important focus of clinical management

Novel human liver-tropic AAV variants define transferable domains that markedly enhance the human tropism of AAV7 and AAV8

Recent clinical successes have intensified interest in using adeno-associated virus (AAV) vectors for therapeutic gene delivery. The liver is a key clinical target, given its critical physiological functions and involvement in a wide range of genetic diseases. Here, we report the bioengineering of a set of next-generation AAV vectors, named AAV-SYDs (where “SYD” stands for Sydney, Australia), with increased human hepato-tropism in a liver xenograft mouse model repopulated with primary human hepatocytes. We followed a two-step process that staggered directed evolution and domain-swapping approaches. Using DNA-family shuffling, we first mapped key AAV capsid regions responsible for efficient human hepatocyte transduction in vivo. Focusing on these regions, we next applied domain-swapping strategies to identify and study key capsid residues that enhance primary human hepatocyte uptake and transgene expression. Our findings underscore the potential of AAV-SYDs as liver gene therapy vectors and provide insights into the mechanism responsible for their enhanced transduction profile

Two rapid assays for screening of patulin biodegradation

Artículo sobre distintos ensayos para comprobar la biodegradación de la patulinaThe mycotoxin patulin is produced by the blue mould pathogen Penicillium expansum in rotting apples during postharvest storage. Patulin is toxic to a wide range of organisms, including humans, animals, fungi and bacteria. Wash water from apple packing and processing houses often harbours patulin and fungal spores, which can contaminate the environment. Ubiquitous epiphytic yeasts, such as Rhodosporidium kratochvilovae strain LS11 which is a biocontrol agent of P. expansum in apples, have the capacity to resist the toxicity of patulin and to biodegrade it. Two non-toxic products are formed. One is desoxypatulinic acid. The aim of the work was to develop rapid, high-throughput bioassays for monitoring patulin degradation in multiple samples. Escherichia coli was highly sensitive to patulin, but insensitive to desoxypatulinic acid. This was utilized to develop a detection test for patulin, replacing time-consuming thin layer chromatography or high-performance liquid chromatography. Two assays for patulin degradation were developed, one in liquid medium and the other in semi-solid medium. Both assays allow the contemporary screening of a large number of samples. The liquid medium assay utilizes 96-well microtiter plates and was optimized for using a minimum of patulin. The semisolid medium assay has the added advantage of slowing down the biodegradation, which allows the study and isolation of transient degradation products. The two assays are complementary and have several areas of utilization, from screening a bank of microorganisms for biodegradation ability to the study of biodegradation pathways

Control of position and movement is simplified by combined muscle spindle and Golgi tendon organ feedback

Whereas muscle spindles play a prominent role in current theories of human motor control, Golgi tendon organs (GTO) and their associated tendons are often neglected. This is surprising since there is ample evidence that both tendons and GTOs contribute importantly to neuromusculoskeletal dynamics. Using detailed musculoskeletal models, we provide evidence that simple feedback using muscle spindles alone results in very poor control of joint position and movement since muscle spindles cannot sense changes in tendon length that occur with changes in muscle force. We propose that a combination of spindle and GTO afferents can provide an estimate of muscle-tendon complex length, which can be effectively used for low-level feedback during both postural and movement tasks. The feasibility of the proposed scheme was tested using detailed musculoskeletal models of the human arm. Responses to transient and static perturbations were simulated using a 1-degree-of-freedom (DOF) model of the arm and showed that the combined feedback enabled the system to respond faster, reach steady state faster, and achieve smaller static position errors. Finally, we incorporated the proposed scheme in an optimally controlled 2-DOF model of the arm for fast point-to-point shoulder and elbow movements. Simulations showed that the proposed feedback could be easily incorporated in the optimal control framework without complicating the computation of the optimal control solution, yet greatly enhancing the system's response to perturbations. The theoretical analyses in this study might furthermore provide insight about the strong physiological couplings found between muscle spindle and GTO afferents in the human nervous system. © 2013 the American Physiological Society

Analysis of factors influencing the ultrasonic fetal weight estimation

Objective: The aim of our study was the evaluation of sonographic fetal weight estimation taking into consideration 9 of the most important factors of influence on the precision of the estimation. Methods: We analyzed 820 singleton pregnancies from 22 to 42 weeks of gestational age. We evaluated 9 different factors that potentially influence the precision of sonographic weight estimation ( time interval between estimation and delivery, experts vs. less experienced investigator, fetal gender, gestational age, fetal weight, maternal BMI, amniotic fluid index, presentation of the fetus, location of the placenta). Finally, we compared the results of the fetal weight estimation of the fetuses with poor scanning conditions to those presenting good scanning conditions. Results: Of the 9 evaluated factors that may influence accuracy of fetal weight estimation, only a short interval between sonographic weight estimation and delivery (0-7 vs. 8-14 days) had a statistically significant impact. Conclusion: Of all known factors of influence, only a time interval of more than 7 days between estimation and delivery had a negative impact on the estimation

Attenuation of Heparan Sulfate Proteoglycan Binding Enhances In Vivo Transduction of Human Primary Hepatocytes with AAV2

Use of the prototypical adeno-associated virus type 2 (AAV2) capsid delivered unexpectedly modest efficacy in an early liver-targeted gene therapy trial for hemophilia B. This result is consistent with subsequent data generated in chimeric mouse-human livers showing that the AAV2 capsid transduces primary human hepatocytes in vivo with low efficiency. In contrast, novel variants generated by directed evolution in the same model, such as AAV-NP59, transduce primary human hepatocytes with high efficiency. While these empirical data have immense translational implications, the mechanisms underpinning this enhanced AAV capsid transduction performance in primary human hepatocytes are yet to be fully elucidated. Remarkably, AAV-NP59 differs from the prototypical AAV2 capsid by only 11 aa and can serve as a tool to study the correlation between capsid sequence/structure and vector function. Using two orthogonal vectorological approaches, we have determined that just 2 of the 11 changes present in AAV-NP59 (T503A and N596D) account for the enhanced transduction performance of this capsid variant in primary human hepatocytes in vivo, an effect that we have associated with attenuation of heparan sulfate proteoglycan (HSPG) binding affinity. In support of this hypothesis, we have identified, using directed evolution, two additional single amino acid substitution AAV2 variants, N496D and N582S, which are highly functional in vivo. Both substitution mutations reduce AAV2's affinity for HSPG. Finally, we have modulated the ability of AAV8, a highly murine-hepatotropic serotype, to interact with HSPG. The results support our hypothesis that enhanced HSPG binding can negatively affect the in vivo function of otherwise strongly hepatotropic variants and that modulation of the interaction with HSPG is critical to ensure maximum efficiency in vivo. The insights gained through this study can have powerful implications for studies into AAV biology and capsid development for preclinical and clinical applications targeting liver and other organs

Giant Electroresistance in Edge Metal-Insulator-Metal Tunnel Junctions Induced by Ferroelectric Fringe Fields

An enormous amount of research activities has been devoted to developing new types of non-volatile memory devices as the potential replacements of current flash memory devices. Theoretical device modeling was performed to demonstrate that a huge change of tunnel resistance in an Edge Metal-Insulator-Metal (EMIM) junction of metal crossbar structure can be induced by the modulation of electric fringe field, associated with the polarization reversal of an underlying ferroelectric layer. It is demonstrated that single three-terminal EMIM/Ferroelectric structure could form an active memory cell without any additional selection devices. This new structure can open up a way of fabricating all-thin-film-based, high-density, high-speed, and low-power non-volatile memory devices that are stackable to realize 3D memory architectureope

Loss of KCNJ10 protein expression abolishes endocochlear potential and causes deafness in Pendred syndrome mouse model

BACKGROUND: Pendred syndrome, a common autosomal-recessive disorder characterized by congenital deafness and goiter, is caused by mutations of SLC26A4, which codes for pendrin. We investigated the relationship between pendrin and deafness using mice that have (Slc26a4(+/+)) or lack a complete Slc26a4 gene (Slc26a4(-/-)). METHODS: Expression of pendrin and other proteins was determined by confocal immunocytochemistry. Expression of mRNA was determined by quantitative RT-PCR. The endocochlear potential and the endolymphatic K(+ )concentration were measured with double-barreled microelectrodes. Currents generated by the stria marginal cells were recorded with a vibrating probe. Tissue masses were evaluated by morphometric distance measurements and pigmentation was quantified by densitometry. RESULTS: Pendrin was found in the cochlea in apical membranes of spiral prominence cells and spindle-shaped cells of stria vascularis, in outer sulcus and root cells. Endolymph volume in Slc26a4(-/- )mice was increased and tissue masses in areas normally occupied by type I and II fibrocytes were reduced. Slc26a4(-/- )mice lacked the endocochlear potential, which is generated across the basal cell barrier by the K(+ )channel KCNJ10 localized in intermediate cells. Stria vascularis was hyperpigmented, suggesting unalleviated free radical damage. The basal cell barrier appeared intact; intermediate cells and KCNJ10 mRNA were present but KCNJ10 protein was absent. Endolymphatic K(+ )concentrations were normal and membrane proteins necessary for K(+ )secretion were present, including the K(+ )channel KCNQ1 and KCNE1, Na(+)/2Cl(-)/K(+ )cotransporter SLC12A2 and the gap junction GJB2. CONCLUSIONS: These observations demonstrate that pendrin dysfunction leads to a loss of KCNJ10 protein expression and a loss of the endocochlear potential, which may be the direct cause of deafness in Pendred syndrome