Ultrasonographic appearance of normal and injured lateral patellar ligaments in the equine stifle.

REASONS FOR PERFORMING STUDY: Ultrasound is widely used in horses with stifle lameness, yet limited information is available regarding the appearance of normal and injured lateral patellar ligaments (LPL). OBJECTIVES: To map the normal ultrasonographic appearance of the LPL. To describe the clinical and ultrasonographic features of LPL injuries. STUDY DESIGN: Descriptive study of healthy horses and retrospective case series. METHODS: Twelve horses without stifle lameness underwent ultrasonographic examination of bilateral LPLs and ultrasonographic features were recorded. Eighteen horses with LPL injury were identified from 1999 to 2011. RESULTS: The normal LPL changes in appearance from origin to insertion. It shows ill-defined margins at the patella, becomes flattened and bilobed over the lateral trochlear ridge, is oval-triangular shaped with variable echogenicity and fibre pattern distal to the LTR, and becomes tapered with striations at the tibial insertion. LPL injury was identified in 18 horses of multiple breeds and uses. All injuries were acute, and 12 had wounds. Eleven horses were severely lame (grade 4-5/5). Ultrasonographic lesions were severe in 78% of cases. The mid to insertional portion of the LPL was most often affected. Radiography showed fractures of the tibial tuberosity (n = 6), patella (n = 4) and lateral trochlear ridge (n = 1). Fractures involved LPL attachments in 9 horses. Five were treated for osteomyelitis and one for synovial sepsis. Recheck ultrasound in 4 horses showed minimal to no change in the appearance of LPL injuries. Nine horses returned to riding, one continued as a broodmare, 2 were retired, one became a broodmare, 2 were lost to follow-up and 3 were subjected to euthanasia owing to concurrent injuries. CONCLUSION: Normal variations in shape, echogenicity and fibre pattern of the LPL are important considerations to prevent false positive diagnoses during ultrasonography. LPL injuries were often severe and associated with craniolateral stifle trauma. Prognosis varied from good to guarded in horses without additional severe injuries.

Kinetics of the translocation and phosphorylation of alphaB-crystallin in mouse heart mitochondria during ex vivo ischemia.

alphaB-crystallin (alphaBC) is a small heat shock protein expressed at high levels in the myocardium where it protects from ischemia-reperfusion damage. Ischemia-reperfusion activates p38 MAP kinase, leading to the phosphorylation of alphaBC on serine 59 (P-alphaBC-S59), enhancing its ability to protect myocardial cells from damage. In the heart, ischemia-reperfusion also causes the translocation of alphaBC from the cytosol to other cellular locations, one of which was recently shown to be mitochondria. However, it is not known whether alphaBC translocates to mitochondria during ischemia-reperfusion, nor is it known whether alphaBC phosphorylation takes place before or after translocation. In the present study, analyses of mitochondrial fractions isolated from mouse hearts subjected to various times of ex vivo ischemia-reperfusion showed that alphaBC translocation to mitochondria was maximal after 20 min of ischemia and then declined steadily during reperfusion. Phosphorylation of mitochondrial alphaBC was maximal after 30 min of ischemia, suggesting that at least in part it occurred after alphaBC association with mitochondria. Consistent with this was the finding that translocation of activated p38 to mitochondria was maximal after only 10 min of ischemia. The overexpression of alphaBC-AAE, which mimics alphaBC phosphorylated on serine 59, has been shown to stabilize mitochondrial membrane potential and to inhibit apoptosis. In the present study, infection of neonatal rat cardiac myocytes with adenovirus-encoded alphaBC-AAE decreased peroxide-induced mitochondrial cytochrome c release. These results suggest that during ischemia alphaBC translocates to mitochondria, where it is phosphorylated and contributes to modulating mitochondrial damage upon reperfusion.

Localization of phosphorylated alphaB-crystallin to heart mitochondria during ischemia-reperfusion.

The cytosolic small heat shock protein alphaB-crystallin (alphaBC) is a molecular chaperone expressed in large quantities in the heart, where it protects from stresses such as ischemia-reperfusion (I/R). Upon I/R, p38 MAP kinase activation leads to phosphorylation of alphaBC on Ser(59) (P-alphaBC-S59), which increases its protective ability. alphaBC confers protection, in part, by interacting with and affecting the functions of key components in stressed cells. We investigated the hypothesis that protection from I/R damage in the heart by P-alphaBC-S59 can be mediated by localization to mitochondria. We found that P-alphaBC-S59 localized to mitochondria isolated from untreated mouse hearts and that this localization increased more than threefold when the hearts were subjected to ex vivo I/R. Mitochondrial P-alphaBC-S59 decreased when hearts were treated with the p38 inhibitor SB-202190. Moreover, SB-202190-treated hearts exhibited more tissue damage and less functional recovery upon reperfusion than controls. I/R activates mitochondrial permeability transition (MPT) pore opening, which increases cell damage. We found that mitochondria incubated with a recombinant mutant form of alphaBC that mimics P-alphaBC-S59 exhibited decreased calcium-induced MPT pore opening. These results indicate that mitochondria may be among the key components in stressed cells with which P-alphaBC-S59 interacts and that this localization may protect the myocardium, in part, by modulating MPT pore opening and, thus, reducing I/R injury.

Response to myocardial ischemia/reperfusion injury involves Bnip3 and autophagy.

Ischemia and reperfusion (I/R) injury is associated with extensive loss of cardiac myocytes. Bnip3 is a mitochondrial pro-apoptotic Bcl-2 protein which is expressed in the adult myocardium. To investigate if Bnip3 plays a role in I/R injury, we generated a TAT-fusion protein encoding the carboxyl terminal transmembrane deletion mutant of Bnip3 (TAT-Bnip3DeltaTM) which has been shown to act as a dominant negative to block Bnip3-induced cell death. Perfusion with TAT-Bnip3DeltaTM conferred protection against I/R injury, improved cardiac function, and protected mitochondrial integrity. Moreover, Bnip3 induced extensive fragmentation of the mitochondrial network and increased autophagy in HL-1 myocytes. 3D rendering of confocal images revealed fragmented mitochondria inside autophagosomes. Enhancement of autophagy by ATG5 protected against Bnip3-mediated cell death, whereas inhibition of autophagy by ATG5K130R enhanced cell death. These results suggest that Bnip3 contributes to I/R injury which triggers a protective stress response with upregulation of autophagy and removal of damaged mitochondria.

Functional and clinical repercussions of myocyte apoptosis in the multifaceted damage by ischemia/reperfusion injury: old and new concepts after 10 years of contributions.

Ten years ago, the first finding of apoptotic cell death on the 'crime scene' of cardiac ischemia/reperfusion injury profoundly dismayed the scientific community. This observation jarred with the deeply rooted conviction that cardiac myocytes stoically 'break, but do not bend' in the fight against ischemia, instead of spontaneously accepting a peaceful demise for the greater good. Ten years later, a number of studies not only proved right the coexistence of necrosis and apoptosis on the ischemic battle field, but also implicated myocyte apoptosis in the pathogenesis of all the shapes and shades that cardiac ischemic injury can take on.

Type II secretory phospholipase A2 binds to ischemic flip-flopped cardiomyocytes and subsequently induces cell death.

Type II secretory phospholipase A2 (sPLA2) is a cardiovascular risk factor. We recently found depositions of sPLA2 in the necrotic center of infarcted human myocardium and normally appearing cardiomyocytes adjacent to the border zone. The consequences of binding of sPLA2 to ischemic cardiomyocytes are not known. To explore a potential effect of sPLA2 on ischemic cardiomyocytes at a cellular level we used an in vitro model. The cardiomyocyte cell line H9c2 or adult cardiomyocytes were isolated from rabbits that were incubated with sPLA2 in the presence of metabolic inhibitors to mimic ischemia-reperfusion conditions. Cell viability was established with the use of annexin V and propidium iodide or 7-aminoactinomycin D. Metabolic inhibition induced an increase of the number of flip-flopped cells, including a population that did not stain with propidium iodide and that was caspase-3 negative. sPLA2 bound to the flip-flopped cells, including those negative for caspase-3. sPLA2 binding induced cell death in these latter cells. In addition, sPLA2 potentiated the binding of C-reactive protein (CRP) to these cells. We conclude that by binding to flip-flopped cardiomyocytes, including those that are caspase-3 negative and presumably reversibly injured, sPLA2 may induce cell death and tag these cells with CRP.

5'-sulfhydryl-modified RNA: initiator synthesis, in vitro transcription, and enzymatic incorporation.

The detailed syntheses of the sulfhydryl-modified guanosine monophosphates 5'-deoxy-5'-thioguanosine-5'-monophosphorothioate (GSMP), O-[omega-sulfhydryl-tetra(ethylene glycol)]-O-(5'-guanosine) monophosphate (5'-HS-PEG4-GMP), and O-[omega-sulfhydryl-di(ethylene glycol)]-O-(5'-guanosine) monophosphate (5'-HS-PEG2-GMP) are reported. Transcription reactions employing GSMP, 5'-HS-PEG4-GMP, or 5'-HS-PEG2-GMP as the initiator nucleotide for T7 RNA polymerase introduce a thiol group at the 5'-end of RNA. The efficiency of thiol incorporation at the 5'-terminus of modified RNA compounds was assayed with three different thiol-reactive biotinylated reagents followed by streptavidin gel-shift methods. The transcription efficiency with various ratios of GTP to 5'-HS-PEG2-GMP was explored by reaction with a sulfhydryl-reactive maleimide-conjugated protein. This is an efficient method to incorporate enzymatically a thiol group into the 5'-end of RNA.

Benign intratesticular cystic lesions: US features.

Benign intratesticular lesions are rare, but recognition is important to avoid unnecessary surgical intervention. The ultrasonographic (US) features that help differentiate benign from malignant intratesticular lesions are emphasized by the authors. Benign lesions include intratesticular simple cysts, tubular ectasia, epidermoid cyst, tunica albuginea cyst, intratesticular varicocele, abscess, and hemorrhage (infarction). US features of cystic malignant neoplasms that help in differentiation of them from benign cystic lesions are also presented. The US appearance of epidermoid cysts varies with the maturation, compactness, and quantity of keratin present. Of the cystic malignant testicular tumors, which can occur anywhere in testicular parenchyma, teratomas are the most frequent to manifest as cystic masses. An abnormal rind of parenchyma with increased echogenicity usually surrounds these lesions. An intratesticular spermatocele communicates with the seminiferous tubules, whereas simple ectasia of the rete testis does not do so directly. These cysts contain spermatozoa and can be septate. The US findings of intratesticular varicocele are similar to those of extratesticular varicocele and include multiple anechoic, serpiginous, tubular structures of varying sizes. Improvements in gray-scale and Doppler US technology allow subtle distinctions between benign and malignant testicular lesions that were not possible a decade earlier.

Phosphorylation of mitochondrial elongation factor Tu in ischemic myocardium: basis for chloramphenicol-mediated cardioprotection.

The objective of this study was to identify the mitochondrial proteins that undergo changes in phosphorylation during global ischemia and reperfusion in the isolated rabbit heart. We also assessed whether the cardioprotective intervention of ischemic preconditioning affected mitochondrial protein phosphorylation. We established a reconstituted system using isolated mitochondria and cytosol from control or ischemic hearts. We found that phosphorylation of a 46-kDa protein on a serine residue was increased in ischemia and that phosphorylation was reduced in control or preconditioned hearts. Using 2D gel electrophoresis and mass spectrometry, we have identified the 46-kDa protein as mitochondrial translational elongation factor Tu (EF-Tu(mt)). These data reveal that ischemia and preconditioning modulate the phosphorylation of EF-Tu(mt) and suggest that the mitochondrial protein synthesis machinery may be regulated by phosphorylation. Phosphorylation of mitochondrial EF-Tu has not been previously described; however, in prokaryotes, EF-Tu phosphorylation inhibits protein translation. We hypothesized that phosphorylation of mitochondrial EF-Tu would inhibit mitochondrial protein translation and attempted to reproduce the effect with inhibition of mitochondrial protein synthesis by chloramphenicol. We found that chloramphenicol pretreatment significantly reduced infarct size, suggesting that mitochondrial protein synthesis is one determinant of myocardial injury during ischemia and reperfusion.

Documenting and comparing medical students' clinical experiences.

CONTEXT: The decentralization of clinical teaching networks over the past decade calls for a systematic way to record the case-mix of patients, the severity of diseases, and the diagnostic procedures that medical students encounter in clinical clerkships. OBJECTIVE: To demonstrate a system that documents medical students' clinical experiences across clerkships. DESIGN AND SETTINGS: Evaluation of a method for recording student-patient clinical encounters using a pocket-sized computer-read patient encounter card at a US university hospital and its 16 teaching affiliates during academic years 1997-1998 through 1999-2000. PARTICIPANTS: A total of 647 third-year medical students who completed patient encounter cards in 3 clerkships: family medicine, pediatrics, and internal medicine. MAIN OUTCOME MEASURES: Number of patient encounters, principal and secondary diagnoses, severity of diseases, and diagnostic procedures as recorded on patient encounter cards; concordance of patient encounter card data with medical records. RESULTS: Students completed 86 011 patient encounter cards: 48 367 cards by 582 students in family medicine, 22 604 cards by 469 students in pediatrics, and 15 040 cards by 531 students in internal medicine. Significant differences were found in students' case-mix of patients, the level of disease severity, and the number of diagnostic procedures performed across the 3 clerkships. Stability of the findings within each clerkship across 3 academic years and the 77% concordance of students' reports of principal diagnosis with faculty's confirmation of diagnosis support the reliability and validity of the findings. CONCLUSIONS: An instrument that facilitates students' documentation of clinical experiences can provide data on important differences among students' clerkship experiences. Data from this instrument can be used to assess the nature of students' clinical education.

Proliferation, not apoptosis, alters epithelial cell migration in small intestine of CFTR null mice.

Expression of a mutated cystic fibrosis transmembrane conductance regulator (CFTR) has been shown to enhance proliferation within CF airways, and cells expressing a mutated CFTR have been shown to be less susceptible to apoptosis. Because the CFTR is expressed in the epithelial cells lining the gastrointestinal tract and all CF mouse models are characterized by gastrointestinal obstruction, we hypothesized that CFTR null mice would have increased epithelial cell proliferation and reduced apoptosis within the small intestine. The rate of intestinal epithelial cell migration from crypt to villus was increased in CFTR null mice relative to mice expressing the wild-type CFTR. This difference in migration could be explained by an increase in epithelial cell proliferation but not by a difference in apoptosis within the crypts of Lieberkühn. In addition, using two independent sets of CF cell lines, we found that epithelial cell susceptibility to apoptosis was unrelated to the presence of a functional CFTR. Thus increased proliferation but not alterations in apoptosis within epithelial cells might contribute to the pathophysiology of CF.

Bid is cleaved by calpain to an active fragment in vitro and during myocardial ischemia/reperfusion.

Reperfusion after myocardial ischemia is associated with a rapid influx of calcium, leading to activation of various enzymes including calpain. Isolated perfused adult rabbit hearts subjected to global ischemia and reperfusion were studied. Calpain or a calpain-like activity was activated within 15 min after reperfusion, and preconditioning suppressed calpain activation. In contrast, caspase activation was not detected although cytochrome c was released after ischemia and reperfusion. The pro-apoptotic BH3-only Bcl-2 family member, Bid, was cleaved during ischemia/reperfusion in the adult rabbit heart. Recombinant Bid was cleaved by calpain to a fragment that was able to mediate cytochrome c release. The calpain cleavage site was mapped to a region within Bid that is extremely susceptible to proteolysis. These findings suggest that there is cross-talk between apoptotic and necrotic pathways in myocardial ischemia/reperfusion injury.

Third-year medical student survey of office preceptorships during the pediatric clerkship.

OBJECTIVE: To assess medical students' interest in a career in pediatrics following their categorical pediatric clerkship. DESIGN: Satisfaction questionnaire to 704 third-year clerks in 5 university medical schools following the pediatric clerkship. METHODS: Analysis of the influence of the community office-based experience compared with the inpatient experience, and examination aspects of the office preceptorship most valued by the medical students. MAIN OUTCOME MEASURE: Satisfaction questionnaire addressing office-based experiences. RESULTS: Third-year pediatric clerks report that the private office setting provides a valuable learning experience, particularly when there is exposure to a wide spectrum of disease and when the preceptor had time to teach. Feelings about pediatrics as career choice rose during the clerkship from neutral to positive, and the frequency of strongly positive feelings rose from 9.2% to 28.6%. In deciding about pediatrics as a career, experiences with patients and residents in the inpatient setting still seem to count more than those experiences in the outpatient setting. CONCLUSION: Categorical pediatric clerkships provide learning environments that influence students positively toward pediatrics as a career choice. This choice is enhanced by encouraging community practitioners with students in their office to expose them to a wide variety of issues and devote time to teaching.

Mitochondria and apoptosis.

Apoptosis is a coordinated sequence of events culminating in the death of the cell. Many of these biochemical processes are regulated by the mitochondria, including the release of proapoptotic molecules in addition to the caspase-activating cofactor, cytochrome c. Pro- and antiapoptotic members of the Bcl-2 family regulate mitochondrial participation in cell death. Current models explaining cytochrome c release are discussed in light of mitochondrial structure and physiology.

Testicular epidermoid cysts: sonographic features with histopathologic correlation.

Testicular epidermoid cysts are rare, accounting for 1% of all testicular tumors. We present the sonographic appearances of epidermoid cysts in 3 cases, together with the histopathologic correlation. In case 1, sonography showed an intratesticular hypoechoic mass with a well-defined echogenic rim; the mass measured 1.8 x 1.5 x 1.5 cm, and there was no evidence of calcification. In case 2, sonography showed a well-circumscribed mass measuring 1.3 x 1.3 x 1.0 cm, with alternating hypoechoic and hyperechoic rings (onion-ring appearance) and no calcifications. In case 3, sonography showed a 2.4- x 2.3- x 2.3-cm, well-circumscribed, oval mass with a heterogeneous echotexture and an outer hypoechoic halo. The mass contained plaque-like regions of increased echogenicity, with peripheral acoustic shadowing from refraction artifact. Hypoechoic clefts were visualized posterior to the plaque-like areas. The triad of findings-sonographic appearance of an onion ring, avascularity on Doppler sonography, and negative results of tumor marker studies-is highly suggestive of an epidermoid cyst.

Pharmacology of caspase inhibitors in rabbit cardiomyocytes subjected to metabolic inhibition and recovery.

Protection of ischemic myocardium is an important unmet need in reperfusion therapy of acute myocardial infarction. Myocardial ischemia and reperfusion induce necrosis and apoptosis in cardiomyocytes. Caspase processing and activation are critical steps in most receptor and nonreceptor pathways of apoptosis. Caspase inhibitors have been shown to reduce ischemia reperfusion injury in cardiac muscle. Information about dose response and time of administration are needed to optimize the design of preclinical studies. We used isolated adult rabbit cardiomyocytes subjected to metabolic inhibition (MI) and recovery to examine the role of caspases and caspase inhibitors, the dose response, and the timing of administration. In vitro inhibitory concentrations (Ki) were determined for purified caspases. Cardiomyocytes subjected to MI were treated with peptidomimetic fluoromethyl ketone inhibitors of caspases before or during MI, or at recovery. Caspase inhibitors were most effective when added before MI and included throughout recovery, but were partially protective if added after MI. The optimal concentration of the inhibitors tested was approximately 10 microM. Protection was sustained when cells were allowed to recover for 4 or 24 h. These results suggest that caspase activation is an important component of myocyte injury mediated by MI and recovery. Low doses of caspase inhibitors were identified that reduce injury in this model system, and further investigations using in vivo models are warranted.

Consultation and therapy in the boarding high school setting.

The tasks of a psychiatric consultant in the boarding high school setting are complex and require a particular set of tools. A thorough familiarity and comfort with the psychology and psychopathology of adolescence are necessities. In my view a psychodynamic point of view is most helpful. I have emphasized that students attending boarding schools are different from all other high school students in that they are experiencing a radical and more or less permanent separation from their homes a full developmental epoch earlier than their nonboarding peers. Whether this separation is helpful and growth promoting or traumatic and disruptive needs to be evaluated for each individual whose family seeks psychiatric opinion about such matters. Broad generalization is not possible. What is required is a careful evaluation of the developmental state of the particular teenager and an assessment of the psychologic meaning of the experience in terms of that child's psychic reality. To this formulation I add that a knowledge of the particular boarding schools under consideration, their cultures, mores, personnel, and administrative and academic styles can be of great additional help in making a decision. In addition to their radical and early separation from home, boarding students find themselves integrated into tightly knit school communities, each with its own unique group and institutional dynamics. The best consultation work will include an understanding of what is occurring at the interface between individual and school community. These interactions can be as critical in determining the fate of a boarding student as can be the interactions between a student living at home and interacting with his or her family. Separated from home and emotionally hungry for new objects to replace the ones left behind, the boarding student may have passionate and intense interactions with the institutions and members of the school community. In this article I have provided a recent example (Ted) of such an intense interaction, albeit one with a terrible and tragic outcome. Such anatomies of suicides as the one I have tried to reconstruct in this case can provide a realistic basis for optimism that a future recurrence can be prevented. Another case (Bob, discussed first) illustrated the potential for profound and far-reaching positive results of the consultation and treatment processes.

Environmentalism unbound: reconnecting and re-envisioning movements for change.

Disputes over environmental discourse have generated divergent pathways and discordant messages over the last three decades. An examination of them becomes a study of environmentalism s roots. The workplace is shown as a hidden and often discounted arena of debate about what constitutes an environmental issue. The triumph of the productionist and limitless consumption views helped to establish a focus on environmental change as a form of consumer action. Since the 1970s though, new forms of environmental discourse and action both community- and production-related have sought to shift the terrain. The possibility of becoming a broader, more socially inclusive movement capable of challenging the very structure and logic of capitalist social order is possible again, including the ability to identify new strategies for action. Overcoming the work/environment divide is perhaps the most contentious question facing the future of the environmental and labor movements. New approaches, including developing a community of interests, revaluing work, and developing an ethic of place (with urban, industrial, and global forms), require that the social and the ecological become joined in the construction of a common vision. When any environmental issue can be seen as socially determined, then environmentalism s great task will be to see itself as a primary agent of social change.

Differential processing of cytosolic and mitochondrial caspases.

The mitochondria have been shown to play a key role in the initiation of caspase activation during apoptosis. Recently, some caspases have been shown to be associated with mitochondria. In this study, we used Jurkat T-lymphoblasts to show that caspases -2 and -3 are located in the mitochondrial intermembrane space, associated with the inner membrane. Caspase-9 is associated with the outer membrane and is exposed to the cytosolic compartment. Caspase activation took place predominantly in the cytosol in response to Fas ligation, but staurosporine treatment led to caspase activation in both cytosol and mitochondria. In response to both Fas and staurosporine treatment, caspase processing could be detected earlier in cytosol than in mitochondria, but this could reflect the limits of sensitive detection by immunoblotting. Only trace amounts of Apaf-1 were found in association with the mitochondria. However, staurosporine treatment led to preferential auto-processing of caspase-9 associated with mitochondria. These findings suggest that mitochondrial caspases are regulated independently of the cytosolic pool of caspases. The data are also consistent with the notion of a caspase nucleation site associated with mitochondria. Using a stable transfected CEM cell line, we show that Bcl-2 suppressed caspase processing in both cytosolic and mitochondrial compartments in response to both staurosporine and Fas ligation.