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Background: Minimally invasive beaded electrosurgical dissectors ("BEED devices") provide simultaneous sharp dissection, blunt dissection, and electrosurgical coagulation while performing 100 cm 2 porcine tissue plane dissections in 0.8 to 3 min with minimal bleeding and no perforations., Objectives: The aim of the study was to report the basic science and potential clinical applications and to video document the speed and quality of planar dissections in in vivo and ex vivo porcine models with thermal damage quantified by thermal and histopathologic measurements. Additionally, in vivo porcine specimens were followed for 90 days to show whether adverse events occurred on a gross or macroscopic basis, as evidenced by photography, videography, physical examination, and dual ultrasonography., Methods: Ex vivo porcine models were subjected to 20, 30, and 50 W in single-stroke passages with BEED dissectors (granted FDA 510(k) clearance (K233002)) with multichannel thermocouple, 3 s delay recordation combined with matching hematoxylin and eosin (H&E) histopathology. In vivo porcine models were subjected to eight 10 × 10 cm dissections in each of 2 subjects at 20, 30, and 50 W and evaluated periodically until 90 days, wherein histopathology for H&E, collagen, and elastin was taken plus standard and Doppler ultrasounds prior to euthanasia., Results: Five to 8 mm width dissectors were passed at 1 to 2 cm/s in ex vivo models (1-10 cm/s in vivo models) with an average temperature rise of 5°C at 50 W. Clinically evidenced seromas occurred in the undressed, unprotected wounds, and resolved well prior to 90 days, as documented by ultrasounds and histopathology., Conclusions: In vivo and ex vivo models demonstrated thermal values that were below levels known to damage subcutaneous adipose tissue or skin. Tissue histopathology confirmed healing parameters while Doppler ultrasound demonstrated normal blood flow in posttreatment tissues., (© The Author(s) 2024. Published by Oxford University Press on behalf of The Aesthetic Society.)

Current risk-adjusted models for donor lung use and lung graft survival do not include donor critical care data. We sought to identify modifiable donor physiologic and mechanical ventilation parameters that predict donor lung use and lung graft survival. This is a prospective observational study of donors after brain death (DBDs) managed by 19 Organ Procurement Organizations from 2016 to 2019. Demographics, mechanical ventilation parameters, and critical care data were recorded at standardized time points during donor management. The lungs were transplanted from 1811 (30%) of 6052 DBDs. Achieving ≥7 critical care endpoints was a positive predictor of donor lung use. After controlling for recipient factors, donor blood pH positively predicted lung graft survival (OR 1.48 per 0.1 unit increase in pH) and the administration of dopamine during donor management negatively predicted lung graft survival (OR 0.19). Tidal volumes ≤8 ml/kg predicted body weight (OR 0.65), and higher positive end-expiratory pressures (OR 0.91 per cm H 2 O) predicted decreased donor lung use without affecting lung graft survival. A randomized clinical trial is needed to inform optimal ventilator management strategies in DBDs., (© 2021 The American Society of Transplantation and the American Society of Transplant Surgeons.)

Background: Recent research shows AKI increases the risk of incident CKD. We hypothesized that perioperative AKI may confer increased risk of subsequent CKD compared to nonperioperative AKI., Methods: A MEDLINE search was performed for "AKI, CKD, chronic renal insufficiency, surgery, and perioperative" and related terms yielded 5209 articles. One thousand sixty-five relevant studies were reviewed. One thousand six were excluded because they were review, animal, or pediatric studies. Fifty-nine studies underwent full manuscript review by two independent evaluators. Seventeen met all inclusion criteria and underwent analysis. Two-by-two tables were constructed from AKI +/- and CKD +/- data. The R package metafor was employed to determine odds ratio (OR), and a random-effects model was used to calculate weighted ORs. Leave-1-out, funnel analysis, and structured analysis were used to estimate effects of study heterogeneity and bias., Results: Nonperioperative studies included studies of oncology, percutaneous coronary intervention, and myocardial infarction patients. Perioperative studies comprised patients from cardiac surgery, vascular surgery, and burns. There was significant heterogeneity, but risk of bias was overall assessed as low. The OR for AKI versus non-AKI patients developing CKD in all studies was 4.31 (95% CI 3.01-6.17; p < 0.01). Nonperioperative subjects demonstrated OR 3.32 for developing CKD compared to non-AKI patients (95% CI 2.06-5.34; p < 0.01) while perioperative patients demonstrated OR 5.20 (95% CI 3.12-8.66; p < 0.01) for the same event., Conclusions: We conclude that studies conducted in perioperative and nonperioperative patient populations suggest similar risk of development of CKD after AKI.

Purpose: Because preterm birth and retinopathy of prematurity (ROP) are associated with poor visual acuity (VA) and altered foveal development, we evaluated relationships among the central retinal photoreceptors, postreceptor retinal neurons, overlying fovea, and VA in ROP., Methods: We obtained optical coherence tomograms (OCTs) in preterm born subjects with no history of ROP (none; n = 61), ROP that resolved spontaneously without treatment (mild; n = 51), and ROP that required treatment by laser ablation of the avascular peripheral retina (severe; n = 22), as well as in term born control subjects (term; n = 111). We obtained foveal shape descriptors, measured central retinal layer thicknesses, and demarcated the anatomic parafovea using automated routines. In subsets of these subjects, we obtained OCTs eccentrically through the pupil (n = 46) to reveal the fiber layer of Henle (FLH) and obtained adaptive optics scanning light ophthalmograms (AO-SLOs) of the parafoveal cones (n = 34) and measured their spacing and distribution., Results: Both VA and foveal depth decreased with increasing ROP severity (term, none, mild, severe). In severe subjects, foveae were broader than normal and the parafovea was significantly enlarged compared to every other group. The FLH was thinner than normal in mild (but not severe) subjects. VA was associated with foveal depth more than group. Density of parafoveal cones did not differ significantly among groups., Conclusions: Foveal structure is associated with loss of VA in ROP. The preserved FLH in severe (relative to mild) eyes suggests treatment may help cone axon development. The significantly larger parafovea and increased outer nuclear layer (ONL) thickness in ROP hint that some developmental process affecting the photoreceptors is not arrested in ROP but rather is supranormal.

Objective: To compare the durations of surgery and anesthesia and the likelihoods of short- and long-term postoperative complications between cats positioned in sternal recumbency versus dorsal recumbency for perineal urethrostomy (PU)., Animals: 247 client-owned cats that underwent PU between January 2004 and December 2015 at 6 veterinary teaching hospitals and 1 private veterinary referral hospital., Procedures: Medical records were reviewed, and signalment, presenting complaints, previous history of urethral obstruction or PU, diet fed, medications administered, indication for PU, durations of surgery and anesthesia for PU, suture type and size, suture pattern for skin closure, and short- and long-term postoperative complications were recorded. Univariable and multivariable analyses were performed to identify differences in durations of surgery and anesthesia and the likelihoods of short- and long-term complications between cats positioned in sternal recumbency and those positioned in dorsal recumbency., Results: Patient position was not associated with durations of surgery and anesthesia for PU, even if a concurrent cystotomy was necessary or the patient required repositioning from sternal to dorsal recumbency. Likewise, patient position was not associated with the likelihood of short- and long-term complications., Conclusions and Clinical Relevance: The observed lack of differences in outcomes between sternal and dorsal recumbency suggested that logistic considerations and personal preference can continue to guide veterinarians when positioning cats for PU.

Calcineurin inhibitors (CNIs) are immunosuppressive drugs used to prevent graft rejection after organ transplant. Common side effects include renal magnesium wasting and hypomagnesemia, which may contribute to new-onset diabetes mellitus, and hypercalciuria, which may contribute to post-transplant osteoporosis. Previous work suggested that CNIs reduce the abundance of key divalent cation transport proteins, expressed along the distal convoluted tubule, causing renal magnesium and calcium wasting. It has not been clear, however, whether these effects are specific for the distal convoluted tubule, and whether these represent off-target toxic drug effects, or result from inhibition of calcineurin. The CNI tacrolimus can inhibit calcineurin only when it binds with the immunophilin, FKBP12; we previously generated mice in which FKBP12 could be deleted along the nephron, to test whether calcineurin inhibition is involved, these mice are normal at baseline. Here, we confirmed that tacrolimus-treated control mice developed hypomagnesemia and urinary calcium wasting, with decreased protein and mRNA abundance of key magnesium and calcium transport proteins (NCX-1 and Calbindin-D 28k ). However, qPCR also showed decreased mRNA expression of NCX-1 and Calbindin-D 28k , and TRPM6. In contrast, KS-FKBP12 -/- mice treated with tacrolimus were completely protected from these effects. These results indicate that tacrolimus affects calcium and magnesium transport along the distal convoluted tubule and strongly suggests that inhibition of the phosphatase, calcineurin, is directly involved., (© 2020 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.)

Osteomyelitis, or bone infection, is often induced by antibiotic resistant Staphylococcus aureus strains of bacteria. Although debridement and long-term administration of antibiotics are the gold standard for osteomyelitis treatment, the increase in prevalence of antibiotic resistant bacterial strains limits the ability of clinicians to effectively treat infection. Bacteriophages (phages), viruses that in a lytic state can effectively kill bacteria, have gained recent attention for their high specificity, abundance in nature, and minimal risk of host toxicity. Previously, we have shown that CRISPR-Cas9 genomic editing techniques could be utilized to expand temperate bacteriophage host range and enhance bactericidal activity through modification of the tail fiber protein. In a dermal infection study, these CRISPR-Cas9 phages reduced bacterial load relative to unmodified phage. Thus we hypothesized this temperate bacteriophage, equipped with the CRISPR-Cas9 bactericidal machinery, would be effective at mitigating infection from a biofilm forming S. aureus strain in vitro and in vivo. In vitro, qualitative fluorescent imaging demonstrated superiority of phage to conventional vancomycin and fosfomycin antibiotics against S. aureus biofilm. Quantitative antibiofilm effects increased over time, at least partially, for all fosfomycin, phage, and fosfomycin-phage (dual) therapeutics delivered via alginate hydrogel. We developed an in vivo rat model of osteomyelitis and soft tissue infection that was reproducible and challenging and enabled longitudinal monitoring of infection progression. Using this model, phage (with and without fosfomycin) delivered via alginate hydrogel were successful in reducing soft tissue infection but not bone infection, based on bacteriological, histological, and scanning electron microscopy analyses. Notably, the efficacy of phage at mitigating soft tissue infection was equal to that of high dose fosfomycin. Future research may utilize this model as a platform for evaluation of therapeutic type and dose, and alternate delivery vehicles for osteomyelitis mitigation., Competing Interests: The authors have declared that no competing interests exist.

In the distal kidney tubule, the steroid hormone aldosterone regulates sodium reabsorption via the epithelial sodium channel (ENaC). Most studies seeking to identify ENaC-regulating aldosterone-induced proteins have used transcriptional profiling of cultured cells. To identify salt-sensitive transcripts in an in vivo model, we used low-NaCl or high-NaCl diet to stimulate or suppress endogenous aldosterone, in combination with magnetic- and fluorescence-activated cell sorting to isolate distal tubule cells from mouse kidney for transcriptional profiling. Of the differentially expressed transcripts, 162 were more abundant in distal tubule cells isolated from mice fed low-NaCl diet, and 161 were more abundant in distal tubule cells isolated from mice fed high-NaCl diet. Enrichment analysis of Gene Ontology biological process terms identified multiple statistically overrepresented pathways among the differentially expressed transcripts that were more abundant in distal tubule cells isolated from mice fed low-NaCl diet, including ion transmembrane transport, regulation of growth, and negative regulation of apoptosis. Analysis of Gene Ontology molecular function terms identified differentially expressed transcription factors, transmembrane transporters, kinases, and G protein-coupled receptors. Finally, comparison with a recently published study of gene expression changes in distal tubule cells in response to administration of aldosterone identified 18 differentially expressed genes in common between the two experiments. When expression of these genes was measured in cortical collecting ducts microdissected from mice fed low-NaCl or high-NaCl diet, eight were differentially expressed. These genes are likely to be regulated directly by aldosterone and may provide insight into aldosterone signaling to ENaC in the distal tubule.

25 years is a relatively short period of time for a medical technology to become a standard of care impacting the treatment of millions of people every year. Yet 25 years ago there were no OCT companies, no OCT products, no OCT markets, and only one journal article published using the term OCT (optical coherence tomography). OCT has had a tremendous scientific, clinical, and economic impact on society. Today, it is estimated that there are ~30 Million OCT imaging procedures performed worldwide every year and the OCT system market is approaching $1B per year. OCT has helped diagnose patients with retinal disease at early treatable stages, preventing or greatly reducing irreversible vision loss. The technology has facilitated pharmaceutical development and contributed to fundamental understanding of disease mechanisms in multiple fields. The invention and translation of OCT from fundamental research to daily clinical practice would not have been possible without a complex ecosystem involving interaction among physics, engineering, and clinical medicine; government funding of fundamental and clinical research; collaborative and competitive research in the academic sector; entrepreneurship and industry; addressing real clinical needs; harnessing the innovation that occurs at the boundaries of disciplines; and economic and societal impact. This invited review paper discusses the translation of OCT from fundamental research to clinical practice and commercial impact, as well as describes the ecosystem that helped power OCT to where it is today and will continue to drive future advances. While OCT is an example of a technology that has had a powerful impact, there are many biomedical technologies which are poised for translation to clinical practice, and it is our hope that highlighting this ecosystem will help accelerate their translation and clinical impact.

Narrowly tuned, selective noise masking of chromatic detection has been taken as evidence for the existence of a large number of color mechanisms (i.e., higher order color mechanisms). Here we replicate earlier observations of selective masking of tests in the (L,M) plane of cone space when the noise is placed near the corners of the detection contour. We used unipolar Gaussian blob tests with three different noise color directions, and we show that there are substantial asymmetries in the detection contours-asymmetries that would have been missed with bipolar tests such as Gabor patches. We develop a new chromatic detection model, which is based on probability summation of linear cone combinations, and incorporates a linear contrast energy versus noise power relationship that predicts how the sensitivity of these mechanisms changes with noise contrast and chromaticity. With only six unipolar color mechanisms (the same number as the cardinal model), the new model accounts for the threshold contours across the different noise conditions, including the asymmetries and the selective effects of the noises. The key for producing selective noise masking in the (L,M) plane is having more than two mechanisms with opposed L- and M-cone inputs, in which case selective masking can be produced without large numbers of color mechanisms.

Purpose: To study the density and packing geometry of the extrafoveal cone photoreceptors in eyes with a history of retinopathy of prematurity (ROP). We used a multimodal combination of adaptive optics (AO) scanning light ophthalmoscopy (SLO) and optical coherence tomography (OCT)., Methods: Cones were identified in subjects (aged 14-26 years) with a history of ROP that was either severe and treated by laser ablation of avascular peripheral retina (TROP; n = 5) or mild and spontaneously resolved, untreated (UROP; n = 5), and in term-born controls (CT; n = 8). The AO-SLO images were obtained at temporal eccentricities 4.5°, 9°, 13.5°, and 18° using both confocal and offset apertures with simultaneous, colocal OCT images. Effects of group, eccentricity, and aperture were evaluated and the modalities compared., Results: In the SLO images, cone density was lower and the packing pattern less regular in TROP, relative to CT and UROP retinae. Although SLO image quality appeared lower in TROP, root mean square (RMS) wavefront error did not differ among the groups. In TROP eyes, cone discrimination was easier in offset aperture images. There was no evidence of cone loss in the TROP OCT images., Conclusions: Low cone density in TROP confocal SLO images may have resulted from lower image quality. Since AO correction in these eyes was equivalent to that of the control group, and OCT imaging showed no significant cone loss, the optical properties of the inner retina or properties of the cones themselves are likely altered in a way that affects photoreceptor imaging.

Case Description: A 4-year-old spayed female Mastiff was evaluated for treatment of chronic nonhealing pressure wounds over both elbow regions resulting from attempts at hypertrophic callus excision., Clinical Findings: The wound bed granulation tissue was mottled red and yellow with hyperemic, rolled epithelial edges. The right wound communicated with a large fluid pocket along the thoracic wall. The dog had an inflammatory leukogram with a left shift., Treatment and Outcome: The wounds were debrided, and tissue specimens were collected for histologic evaluation, microbial culture, and bacterial identification by means of molecular diagnostic techniques. The left wound was closed immediately. Calcium alginate rope with silver was packed into the right wound. Vacuum-assisted closure was applied for 6 days. Debridement was repeated, and a thoracodorsal axial pattern flap was used to cover the wound. Systemic treatment with antimicrobials was initiated, and pressure over the elbow regions was relieved. Bacterial biofilms were identified histologically in tissue specimens from both wounds. Staphylococcus intermedius, Staphylococcus epidermidis, and Streptococcus canis were cultured and identified by 16S rRNA fragment sequencing. Pyrosequencing identified multiple bacterial species and no fungal organisms. Both wounds healed successfully., Clinical Relevance: Biofilms are implicated in infected orthopedic implants in veterinary patients; however, this is the first report of a bacterial biofilm in chronic wounds in a dog. In human wound care, extensive debridement is performed to disrupt the biofilm; a multimodal treatment approach is recommended to delay reformation and help clear the infection. In this case, biofilm reformation was prevented by systemic treatment with antimicrobials, by reducing local pressure on the wounds, and by wound closure.

We present a numerical approach to extract the dispersion mismatch in ultrahigh-resolution Fourier domain optical coherence tomography (OCT) imaging of the retina. The method draws upon an analogy with a Shack-Hartmann wavefront sensor. By exploiting mathematical similarities between the expressions for aberration in optical imaging and dispersion mismatch in spectral / Fourier domain OCT, Shack-Hartmann principles can be extended from the two-dimensional paraxial wavevector space (or the x-y plane in the spatial domain) to the one-dimensional wavenumber space (or the z-axis in the spatial domain). For OCT imaging of the retina, different retinal layers, such as the retinal nerve fiber layer (RNFL), the photoreceptor inner and outer segment junction (IS/OS), or all the retinal layers near the retinal pigment epithelium (RPE) can be used as point source beacons in the axial direction, analogous to point source beacons used in conventional two-dimensional Shack-Hartman wavefront sensors for aberration characterization. Subtleties regarding speckle phenomena in optical imaging, which affect the Shack-Hartmann wavefront sensor used in adaptive optics, also occur analogously in this application. Using this approach and carefully suppressing speckle, the dispersion mismatch in spectral / Fourier domain OCT retinal imaging can be successfully extracted numerically and used for numerical dispersion compensation to generate sharper, ultrahigh-resolution OCT images.

Proper regulation of the Wingless/Wnt signaling pathway is essential for normal development. The scaffolding protein Axin plays a key role in this process through interactions with Drosophila Shaggy and Armadillo. In the current studies, we used a yeast two-hybrid assay to identify ten amino acids in Axin that are critical for in vitro interaction with Shaggy and two for interaction with Armadillo. We then generated five Axin variants in which individual putative contact amino acids were mutated and compared their activity, as assayed by rescue of axin null mutant flies, to that of Axin lacking the entire Shaggy (AxinDeltaSgg) or Armadillo (AxinDeltaArm) binding domain. Although we expected these mutants to function identically to Axin in which the entire binding domain was deleted, we instead observed a spectrum of phenotypic rescue. Specifically, two point mutants within the Shaggy binding domain showed loss of activity similar to that of AxinDeltaSgg and dominantly interfered with complex function, whereas a third mutant allele, AxinK446E, retained most function. Two Axin point mutants within the Armadillo binding domain were weak alleles and retained most function. These findings demonstrate the importance of in vivo verification of the role of specific amino acids within a protein.

Background: Autosomal recessive primary microcephaly (MCPH) is a model disease to study human neurogenesis. In affected individuals the brain grows at a reduced rate during fetal life resulting in a small but structurally normal brain and mental retardation. The condition is genetically heterogeneous with mutations in ASPM being most commonly reported., Methods and Results: We have examined this further by studying three cohorts of microcephalic children to extend both the phenotype and the mutation spectrum. Firstly, in 99 consecutively ascertained consanguineous families with a strict diagnosis of MCPH, 41 (41%) were homozygous at the MCPH5 locus and all but two families had mutations. Thus, 39% of consanguineous MCPH families had homozygous ASPM mutations. Secondly, in 27 non-consanguineous, predominantly Caucasian families with a strict diagnosis of MCPH, 11 (40%) had ASPM mutations. Thirdly, in 45 families with a less restricted phenotype including microcephaly and mental retardation, but regardless of other neurological features, only 3 (7%) had an ASPM mutation. This report contains 27 novel mutations and almost doubles the number of MCPH associated ASPM mutations known to 57. All but one of the mutations lead to the use of a premature termination codon, 23 were nonsense mutations, 28 deletions or insertions, 5 splicing, and 1 was a translocation. Seventeen of the 57 mutations were recurrent. There were no definitive missense mutations found nor was there any mutation/phenotype correlation. ASPM mutations were found in all ethnic groups studied., Conclusion: This study confirms that mutations in ASPM are the most common cause of MCPH, that ASPM mutations are restricted to individuals with an MCPH phenotype, and that ASPM testing in primary microcephaly is clinically useful.

Deletions of chromosome 1q42-q44 have been reported in a variety of developmental abnormalities of the brain, including microcephaly (MIC) and agenesis of the corpus callosum (ACC). Here, we describe detailed mapping studies of patients with unbalanced structural rearrangements of distal 1q4. These define a 3.5-Mb critical region extending from RP11-80B9 to RP11-241M7 that we hypothesize contains one or more genes that lead to MIC and ACC when present in only one functional copy. Next, mapping of a balanced reciprocal t(1;13)(q44;q32) translocation in a patient with postnatal MIC and ACC demonstrated a breakpoint within this region that is situated 20 kb upstream of AKT3, a serine-threonine kinase. The murine orthologue Akt3 is required for the developmental regulation of normal brain size and callosal development. Whereas sequencing of AKT3 in a panel of 45 patients with ACC did not demonstrate any pathogenic variations, whole-mount in situ hybridization confirmed expression of Akt3 in the developing central nervous system during mouse embryogenesis. AKT3 represents an excellent candidate for developmental human MIC and ACC, and we suggest that haploinsufficiency causes both postnatal MIC and ACC.

Pressure overload-induced cardiac hypertrophy leads to decreased contractile performance, frequently progressing to heart failure. Cardiac hypertrophy and heart failure can be accompanied by the so-called sick thyroid syndrome, resulting in decreased serum T(3) levels along with decreased expression of thyroid hormone receptors (TRalpha1 and TRbeta1) and sarco(endo)plasmic reticulum Ca-ATPase (SERCA). Because the binding of T(3) occupied receptors to the thyroid response elements in the SERCA promotor can increase gene expression, we wanted to determine whether increasing TR expression in the hypertrophied heart could also improve SERCA expression and cardiac function. Mice subjected to aortic constriction to generate pressure overload-induced hypertrophy were also subjected to gene therapy using adeno-associated virus (AAV) expressing either TRalpha1 or TRbeta1, with LacZ expressing AAV serving as control. After 8 wk of aortic constriction, a similar degree of hypertrophy was observed in all three groups; however, mice treated with TRalpha1 or TRbeta1 showed improved contractile function. Administration of a physiological dose of T(3) increased serum T(3) levels only into the lower range of normal. This T(3) dose, with or without AAV TR treatment, did not result in any significant increase in contractile performance. Calcium transients measured in isolated myocytes also exhibited an enhanced rate of decay associated with TRalpha1 or TRbeta1 treatment. Western blot analysis showed increased SERCA expression in the TRalpha1- or TRbeta1-treated groups relative to the LacZ-treated control group. These results demonstrate that increasing TR expression in the hypertrophied heart is associated with an improvement in contractile function and increased SERCA expression.

Inducible heat shock protein 70 (HSP70i) has been shown to exert a protective effect in hearts subjected to ischemia-reperfusion. Although studied in heat-shocked animals and in transgenic mice that constitutively overexpress the protein, the therapeutic application of the protein in the form of a viral vector-mediated HSP70i expression has not been widely examined. Accordingly, we have examined the effects of HSP70i delivered in vivo to the left ventricular free wall of the heart via viral gene therapy in mice. The affect of virally mediated HSP70i expression in preserving cardiac function following ischemia-reperfusion was examined after short-term expression (5-day adenovirus mediated) and long-term expression (8-mo adeno-associated virus mediated) in mice by subjecting ex vivo Langendorff perfused hearts to a regime of ischemia-reperfusion. Both vectors were capable of increasing HSP70i expression in the heart, and neither vector had any effect on cardiac function during aerobic (preischemic) perfusion when compared with corresponding controls. In contrast, both adenovirus-mediated and adeno-associated virus-mediated expression of HSP70i improved the contractile recovery of the heart after 120 min of reperfusion following ischemia. This study demonstrates the feasibility of using both short- and long-term expression of virally mediated HSP70i as a therapeutic intervention against cardiac ischemia-reperfusion injury.

T3 regulates transcription of the rat sarcoendoplasmic reticulum calcium ATPase in the heart. The T3 effect is mediated by three differently configured T3 response elements (TREs). Here we report the mutation of each individual TRE in the promoter and the contribution of each TRE on gene expression. Mutation of TRE1, a direct repeat element, exerted the strongest T3 response, compared with TRE2 and TRE3, which are inverted palindromes. The isolated TRE2 and TRE3, which showed no response (TRE2) or were weakly positive with T3 (TRE3), became strong negative regulatory elements with the T3 analog GC-1. We found that TRE1 recruits corepressor complexes containing nuclear receptor corepressor and histone deacetylase 3 in the absence of ligand, and steroid receptor coactivator-1-containing coactivator complexes with both T3 and GC-1. TRE3 bound the same corepressor complexes without ligand but showed only a weak association with steroid receptor coactivator-1 with T3 and a strong association with corepressor complexes with GC-1. Thus, GC-1 appears to control cofactor association differentially on these two sarcoendoplasmic reticulum calcium ATPase TREs, which could be the mechanism of ligand-dependent transcriptional activation and repression observed with the isolated TRE1 and TRE3 elements. Because the x-ray crystal structures of GC-1 and T3 complexed with the TR ligand binding domain are superimposable, the results imply that GC-1 and T3 induce differential effects on the receptor that are not evident in the static structures but must occur in the dynamic setting of receptor function. These results have implications for selective modulation of receptor function by agonist ligands.

Although it is known that insulin-dependent (type 1) diabetes results in depressed contractile performance associated with diminished sarcoendoplasmic reticular Ca2+-ATPase (SERCA2a) activity, findings in insulin-resistant (type 2) diabetes suggest a less clear association. The db/db insulin-resistant mouse model exhibits decreased cardiac performance both in situ and in isolated ex vivo working hearts. In this study, contractile performance and calcium transients were measured in Langendorff-perfused hearts and isolated cardiac myocytes. Diabetic (db/db) mouse hearts demonstrated decreased rates of contraction, relaxation, and pressure development. Calcium transients from isolated myocytes revealed significantly lower diastolic and systolic levels of calcium in diabetic hearts. Furthermore, the decay rate of the calcium transient was significantly reduced in diabetic myocytes, suggesting a diminished capacity for cytosolic calcium removal not associated with a change in sodium-calcium exchanger activity. Calcium leakage from the sarcoplasmic reticulum (SR) measured using tetracaine was significantly increased in diabetic myocytes. Western blot analysis indicated only a small decrease in SERCA2a expression in diabetic mice, but a large increase in phospholamban expression. Expression of the ryanodine receptor did not differ between groups. In conclusion, the decreased contractile function observed in the db/db diabetic mouse model appears to be related to decreased calcium handling by the SR.

Thyroid hormone (T3) influences cardiac function, and mice with deletion of thyroid hormone receptor (TR)alpha have diminished cardiac function. TR alpha 1 represents 70% and TR beta 1 represents the remaining 30% of TR in ventricular myocytes, and its role in cardiac function is not well established. To determine the role of TR beta 1 in detail, we compared contractility in isolated perfused hearts from wild-type (WT) and TR beta knockout mice under normal and increased work load. TR beta knockout hearts showed contractile function similar to WT hearts at baseline and under conditions of enhanced demand. To gain insight into the role of TR beta, we used mice with a homozygous mutation in exon 10 of TR beta encoding the dominant negative PV mutant (TR beta PV) expressed from the endogenous TR beta promoter. TR beta PV mice treated with 6-propyl-2-thiouracil and supplemented with T3 to make them euthyroid have decreased contractility with negative and positive rates of relaxation and contraction as well as peak systolic pressure diminished by 35 +/- 5, 34 +/- 6, and 35 +/- 6% in comparison with WT mice. Heart rate is diminished by 36 +/- 7%, which is accompanied by decreased expression of the pacemaker-related gene hyperpolarization-activated cyclic nucleotide-gated 4 (HCN4). The expression of TR beta 1 in the pacemaker myocytes of the sinoatrial node was confirmed by quantitation of TR alpha 1 and TR beta 1 mRNA in sinoatrial node, which showed that TR beta 1 mRNA represents 27.5 +/- 1.6% of the ligand-binding isoforms of the TR. In summary, although TR beta is expressed at much lower levels in all regions of the heart than TR alpha 1, expression of the strong dominant negative TR beta PV mutant results in decreased contractile function and heart rate.

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Objective: Prolonged cardiac contraction and relaxation in hypothyroidism are in part related to diminished expression of the gene coding for the calcium pump of the sarcoplasmic reticulum (SERCA2a). Therefore, we examined whether or not transgenic SERCA2a gene expression in mice may compensate for the cardiac effects of hypothyroidism., Methods: SERCA2a mRNA and protein were analyzed from hearts of euthyroid and hypothyroid mice of wild-type or SERCA2a transgene status. Contractile function was studied in isolated left ventricular papillary muscles., Results: We found significant decreases of SERCA2a mRNA and protein levels in hearts of hypothyroid wild-type mice in comparison with euthyroid wild-type mice (controls). Papillary muscles from hypothyroid wild-type mice showed significant increases in time to peak contraction and relaxation times compared with controls. In contrast, SERCA2a mRNA and protein levels were significantly higher in hypothyroid SERCA2a transgenic mice than in hypothyroid wild-type mice. The transgene led to a functional improvement by compensating for the prolonged contraction and relaxation of papillary muscles., Conclusions: Our murine model of hypothyroidism revealed decreases in SERCA2a gene expression accompanied by prolonged contraction and relaxation of papillary muscles, and an improvement of the contractile phenotype due to compensated SERCA2a gene expression in SERCA2a transgenic mice.

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The heart has been recognized as a major target of thyroid hormone action. Our study investigates both the regulation of cardiac-specific genes and contractile behavior of the heart in the presence of a mutant thyroid hormone receptor beta1 (T3Rbeta1-delta337T) derived from the S kindred. The mutant receptor was originally identified in a patient with generalized resistance to thyroid hormone. Cardiac expression of the mutant receptor was achieved by a transgenic approach in mice. As the genes for myosin heavy chains (MHC alpha and MHC beta) and the cardiac sarcoplasmic reticulum Ca2+ adenosine triphosphatase (SERCA2) are known to be regulated by T3, their cardiac expression was analyzed. The messenger RNA levels for MHC alpha and SERCA2 were markedly down-regulated, MHC beta messenger RNA was up-regulated. Although T3 levels were normal in these animals, this pattern of cardiac gene expression mimics a hypothyroid phenotype. Cardiac muscle contraction was significantly prolonged in papillary muscles from transgenic mice. The electrocardiogram of transgenic mice showed a substantial prolongation of the QRS interval. Changes in cardiac gene expression, cardiac muscle contractility, and electrocardiogram are compatible with a hypothyroid cardiac phenotype despite normal T3 levels, indicating a dominant negative effect of the T3Rbeta mutant.

Background: OCT can image plaque microstructure at a level of resolution not previously demonstrated with other imaging techniques because it uses infrared light rather than acoustic waves., Objectives: To compare optical coherence tomography (OCT) and intravascular ultrasound (IVUS) imaging of in vitro atherosclerotic plaques., Methods: Segments of abdominal aorta were obtained immediately before postmortem examination. Images of 20 sites from five patients were acquired with OCT (operating at an optical wavelength of 1300 nm which was delivered to the sample through an optical fibre) and a 30 MHz ultrasonic transducer. After imaging, the microstructure of the tissue was assessed by routine histological processing., Results: OCT yielded superior structural information in all plaques examined. The mean (SEM) axial resolution of OCT and IVUS imaging was 16 (1) and 110 (7), respectively, as determined by the point spread function from a mirror. Furthermore, the dynamic range of OCT was 109 dB compared with 43 dB for IVUS imaging., Conclusions: OCT represents a promising new technology for intracoronary imaging because of its high resolution, broad dynamic range, and ability to be delivered through intravascular catheters.

Background: Optical coherence tomography (OCT) is an recently developed medical diagnostic technology that uses back-reflected infrared light to perform in situ micron scale tomographic imaging. In this work, we investigate the ability of OCT to perform micron scale tomographic imaging of the internal microstructure of in vitro atherosclerotic plaques., Methods and Results: Aorta and relevant nonvascular tissue were obtained at autopsy. Two-dimensional cross-sectional imaging of the exposed surface of the arterial segments was performed in vitro with OCT. A 1300-nm wavelength, superluminescent diode light source was used that allows an axial spatial resolution of 20 microns. The signal-to-noise ratio was 109 dB. Images were displayed in gray scale or false color, Imaging was performed over 1.5 mm into heavily calcified tissue, and a high contrast was noted between lipid- and water-based constituents, making OCT attractive for intracoronary imaging. The 20-microns axial resolution of OCT allowed small structural details such as the width of intimal caps and the presence of fissures to be determined. The extent of lipid collections, which had a low backscattering intensity, also were well documented., Conclusions: OCT represents a promising new technology for imaging vascular microstructure with a level of resolution not previously achieved with the use of other imaging modalities. It does not required direct contact with the vessel wall and can be performed with a catheter integrated with a relatively inexpensive optical fiber. The high contrast among tissue constituents, high resolution, and ability to penetrate heavily calcified tissue make OCT an attractive new imaging technology for intracoronary diagnostics.

Optical coherence tomography is a new imaging technique that can perform high-resolution, micrometre-scale, cross-sectional imaging in biological systems. The technology has been developed, and reduced to, preliminary clinical practice in ophthalmology. The challenging problem that OCT may address is the development of 'optical biopsy' techniques. These techniques can provide diagnostic imaging of tissue morphology without the need for excision of specimens. Many investigations remain to identify optimal areas for clinical application, and additional engineering must be done to integrate vertically the technology and to reduce it to clinical practice. Nevertheless, preliminary studies indicate the feasibility of developing this technology for a wide range of clinical and research diagnostic imaging applications. The ability to non-excisionally evaluate tissue morphology using a catheter or an endoscope could have a significant impact on the diagnosis and management of a wide range of diseases.

A technique called optical coherence tomography (OCT) has been developed for noninvasive cross-sectional imaging in biological systems. OCT uses low-coherence interferometry to produce a two-dimensional image of optical scattering from internal tissue microstructures in a way that is analogous to ultrasonic pulse-echo imaging. OCT has longitudinal and lateral spatial resolutions of a few micrometers and can detect reflected signals as small as approximately 10(-10) of the incident optical power. Tomographic imaging is demonstrated in vitro in the peripapillary area of the retina and in the coronary artery, two clinically relevant examples that are representative of transparent and turbid media, respectively.

Design considerations for a heterodyne spatial tracking system utilizing pupil plane processing techniques and its advantages over traditional focal plane processing are described. Noise performance bounds, optimal and suboptimal local oscillator distributions, pull-in performance, and applications other than spatial tracking are discussed. Experimental verification of a one-axis closed-loop tracking system is presented.