Mice with mutations in Mahogunin ring finger-1 (Mgrn1) exhibit abnormal patterning of the left-right axis.

Mahogunin Ring Finger 1 (Mgrn1) encodes a RING-containing protein with ubiquitin ligase activity that has been implicated in pigment-type switching. In addition to having dark fur, mice lacking MGRN1 develop adult-onset spongy degeneration of the central nervous system and have reduced embryonic viability. Observation of complete situs inversus in a small proportion of adult Mgrn1 mutant mice suggested that embryonic lethality resulted from congenital heart defects due to defective establishment and/or maintenance of the left-right (LR) axis. Here we report that Mgrn1 is expressed in a pattern consistent with a role in LR patterning during early development and that many Mgrn1 mutant embryos show abnormal expression of asymmetrically expressed genes involved in LR patterning. A range of complex heart defects was observed in 20-25% of mid-to-late gestation Mgrn1 mutant embryos and another 20% were dead. This finding was consistent with 46-60% mortality of mutants by weaning age. Our results indicate that Mgrn1 acts early in the LR signaling cascade and is likely to provide new insight into this developmental process as Nodal expression was uncoupled from expression of other Nodal-responsive genes in Mgrn1 mutant embryos. Our work identifies a novel role for MGRN1 in embryonic patterning and suggests that the ubiquitination of MGRN1 target genes is essential for the proper establishment and/or maintenance of the LR axis.     

An enzyme‐linked immunosorbent assay to detect the presence of paralytic shellfish poison using an induced crab protein marker

A purified, high molecular weight protein (referred to as saxitoxin‐induced protein, SIP), was obtained from crabs, Hemigrapsus oregonesis, by affinity chromatography prior to use in a homologous crab SIP enzyme‐linked immunosorbent assay (ELISA) procedure. The SIP measured in H. oregonesis control crabs given acute saxitoxin (SAX) challenge injections (SAX range 0–50 ng), was less than the amount of SIP present in H. oregonesis crabs exposed to a natural toxic dinoflagellate outbreak. The latter were collected from a paralytic shellfish poison (PSP) contaminated coastal area which also contained PSP toxic butterclams (2000 μg PSP per 100 g shellfish), tested by the conventional mouse lethality bioassay procedure. These ELISA results were confirmed by an immunoblotting procedure using anti‐SIP antibody. An immunoblotting procedure of purified SIP and crude SIP antiserum revealed no cross‐reactivity with control, SAX uninjected crabs, thereby indicating specificity of the assay. The method is fast and useful for the screening of antigens expressed in crabs as a consequence of PSP, and represents a procedure that will complement the standard mouse bioassay.     

Germinal center formation in mice lacking αβ T cells

T cells are essential for inducing clonal B cell expansion in germinal centers during T cell-dependent antibody responses. However, class-switched antibodies are readily detectable in TCRα-deficient mice that congenitally lack αβ T cells, including those such as IgG1 that are considered to be dependent on collaboration between B cells and αβ T cells. This observation suggests that a novel form of B:T collaboration may be evident in TCRα^?/? mice. We report that germinal centers develop spontaneously in mice lacking T cell receptor α genes (TCRα^?/?), despite the absence of αβ T cells. They are not seen in TCRβ^?/? mice kept in similar conditions. Both strains of mice have γδ T cells, but it is a subset of T cells expressing TCRβ and CD4 that is dominant in the germinal centers of TCRα^?/? mice. Exceptionally, germinal centers were associated with CD4⁺ γδ T cells. The expression of CD4 seems to be important, for few extrafollicular T cells have CD4 and CD4 is largely absent from TCRβ^?/? T cells. The CD4⁺ TCRβ cells may help B cells produce autoantibodies that have been identified in TCRα^?/? mice.     

The role of size, sequence and haplotype in the stability of FRAXA and FRAXE alleles during transmission

Factors involved in the stability of trinucleotide repeats during transmission were studied in 139 families in which a full mutation, premutation or intermediate allele at either FRAXA or FRAXE was segregating. The transmission of alleles at FRAXA, FRAXE and four microsatellite loci were recorded for all individuals. Instability within the minimal and common ranges (0-40 repeats for FRAXA, 0-30 repeats for FRAXE) was extremely rare; only one example was observed, an increased in size at FRAXA from 29 to 39 repeats. Four FRAXA and three FRAXE alleles in the intermediate range (41-60) repeats for FRAXA, 31-60 for FRAXE) were unstably transmitted. Instability was more frequent for FRAXA intermediate alleles that had a tract of pure CGG greater than 37 although instability only occurred in two of 13 such transmissions: the changes observed were limited to only one or two repeats. Premutation FRAXA alleles over 100 repeats expanded to a full mutation during female transmission in 100% of cases, in agreement with other published series. There was no clear correlation between haplotype and probability of expansion of FRAXA premutations. Instability at FRAXA or FRAXE was more often observed in conjunction with a second instability at an independent locus suggesting genomic instability as a possible mechanism by which at least some FRAXA and FRAXE mutations arise.      

Effect of the AirStirrup in Controlling Ankle Inversion Stress

Subtalar ankle inversion with and without AirStirrup application was evaluated using high speed cinematographic techniques and a specially designed platform that inverts the ankle 35 O. Eighteen subjects aged 19 to 35 and with no history of ankle injury participated in the study. Two trials were filmed for each subject. Although the same ankle was tested in each trial, AirStirrups were applied to both ankles in one trial and to neither in the other trial so subjects could not anticipate the inversion stress. Points marked on the knee (posterior), the Achilles tendon, and the distal calcaneous were digitized. These data were smoothed employing cubic splines and used to calculate the maximum angular displacement (inversion) at the subtalar joint in each of the conditions tested. A t-test revealed a significantly (p c 0.001) larger inversion angle for the ankle not braced with fhe AirStirrup. J Orthop Sports Phys Ther 1987;9(5):190-193.     

Market model addresses hospitals' need for decision support

A Decision Support System should be approached, not as “starting over,” but as a natural extension of the design and development of a hospital's current HIS. Integrated, real-time HISs generate reams of information that, when combined with relevant external data, provide the essential information base for a hospital DDS. The Travenol Market Model is discussed as an example of a DSS specific to hospital needs.     

Recent advances in pathology as applied to orbital biopsy. Practical considerations

Advances in pathology allow for more specific diagnoses of orbital disease. The authors discuss the value of awareness of advances in cytology, histochemistry, immunohistochemistry and electron microscopy as applied to orbital disease. Modern cytologic technique can aid in clearer visualization of cellular detail with improved diagnosis of thin needle aspiration biopsies. Histochemistry offers an increasing range of methods for identification of cellular and extracellular substances such as amyloid, fibrin, neuroglia, and collagen. Immunohistochemistry allows for identification of an ever-increasing number of component antigens including immunoglobulins, myoglobins, keratin, glial fibre protein, etc. Electronmicroscopic technique including plastic embedding allow for specific identification of lesions based on subcellular components and characteristic nuclear, cytoplasmic, membrane, basement membrane and stromal components. The value of these methods has been demonstrated with case presentations of "small round cell tumors" of the adult and child. In addition, the pathologic diagnosis of several rare lesions of the orbit including neuroendocrine carcinoma, histiocytosis X, simultaneously occurring poorly differentiated mucoepidermoid carcinoma of lacrimal gland and adenocarcinoma of the prostate are demonstrated to underline the advances in technology. Emphasis is placed on the management of biopsy material to maximize diagnostic potential.     

Acquired Idiopathic Stiffness After Contemporary Total Knee Arthroplasty: Incidence,Risk Factors,and Results Over 25 Years

BackgroundAcquired idiopathic stiffness (AIS) remains a common failure mode of contemporary total knee arthroplasties (TKAs). The present study investigated the incidence of AIS and manipulation under anesthesia (MUA) at a single institution over time, determined outcomes of MUAs, and identified risk factors associated with AIS and MUA.MethodsWe identified 9771 patients (12,735 knees) who underwent primary TKAs with cemented, modular metal-backed, posterior-stabilized implants from 2000 to 2016 using our institutional total joint registry. Mean age was 68 years, 57% were female, and mean body mass index was 33 kg/m². Demographic, surgical, and comorbidity data were investigated via univariate Cox proportional hazard models and fit to an adjusted multivariate model to access risk for AIS and MUA. Mean follow-up was 7 years.ResultsDuring the study period, 456 knees (3.6%) developed AIS and 336 knees (2.6%) underwent MUA. Range of motion (ROM) increased a mean of 34° after the MUA; however, ROM for patients treated with MUA was inferior to patients without AIS at final follow-up (102° vs 116°, P < .0001). Significant risk factors included younger age (HR 2.3, P < .001), increased tourniquet time (HR 1.01, P < .001), general anesthesia (HR 1.3, P = .007), and diabetes (HR 1.5, P = .001).ConclusionAcquired idiopathic stiffness has continued to have an important adverse impact on the outcomes of a subset of patients undergoing primary TKAs. When utilized, MUA improved mean ROM by 34°, but patients treated with MUA still had decreased ROM compared to patients without AIS. Importantly, we identified several significant risk factors associated with AIS and subsequent MUA.Level of EvidenceLevel III, retrospective comparative study.     

A phase 3, randomized,double-blind,parallel-group study to evaluate tezacaftor/ivacaftor in people with cystic fibrosis heterozygous for F508del-CFTR and a gating mutation

BackgroundTezacaftor (TEZ)/ivacaftor (IVA) is an approved CFTR modulator shown to be efficacious and generally safe and well tolerated in people ≥12 years of age with cystic fibrosis (CF) homozygous for the F508del-CFTR mutation or heterozygous for the F508del-CFTR mutation and a residual function mutation. Although previous studies with IVA alone showed clinical benefits in people with CFTR gating mutations, TEZ/IVA has not yet been evaluated in a Phase 3 study of participants heterozygous for F508del-CFTR and a gating mutation (F/gating genotypes). Here, we present results from a randomized, double-blind, IVA-controlled, parallel-group, Phase 3 study assessing the efficacy, safety, and pharmacokinetics (PK) of TEZ/IVA in participants ≥12 years of age with F/gating genotypes.MethodsEnrolled participants entered a 4-week IVA run-in period to create a stable IVA baseline. Participants were then randomized to receive IVA or TEZ/IVA for 8 weeks in an active comparator treatment period (ACTP). The primary endpoint was absolute change in percent predicted forced expiratory volume in 1 second (ppFEV₁). Key secondary endpoints were relative change in ppFEV₁ and absolute change in CF Questionnaire–Revised respiratory domain score. Secondary endpoints included absolute change in sweat chloride (SwCl) concentration, PK parameters, and safety. All endpoints except PK parameters and safety were assessed from baseline through Week 8.ResultsSixty-nine participants (92.0%) in the IVA group and 75 participants (98.7%) in the TEZ/IVA group completed treatment. No improvements were seen in efficacy endpoints from baseline at the end of the IVA run-in period through the end of the ACTP in the IVA group. No significant differences in ppFEV₁ or any key secondary endpoint were observed between the IVA and TEZ/IVA groups. SwCl concentrations decreased more in the TEZ/IVA versus IVA group during the ACTP. The safety profile and PK parameters of TEZ/IVA were consistent with those of previous studies in participants ≥12 years of age with CF.ConclusionsThis Phase 3 study showed that the dual-combination regimen of TEZ/IVA demonstrated clinical efficacy but did not have significantly greater clinical efficacy than IVA alone in participants ≥12 years of age with F/gating genotypes. However, as reported in other studies, TEZ/IVA was generally safe and well tolerated (NCT02412111).     

Photothermal effects of laser tissue soldering

Low-strength anastomoses and thermal damage of tissue are major concerns in laser tissue welding techniques where laser energy is used to induce thermal changes in the molecular structure of the tissues being joined, hence allowing them to bond together. Laser tissue soldering, on the other hand, is a bonding technique in which a protein solder is applied to the tissue surfaces to be joined, and laser energy is used to bond the solder to the tissue surfaces. The addition of protein solders to augment tissue repair procedures significantly reduces the problems of low strength and thermal damage associated with laser tissue welding techniques. Investigations were conducted to determine optimal solder and laser parameters for tissue repair in terms of tensile strength, temperature rise and damage and the microscopic nature of the bonds formed. An in vitro study was performed using an 808 nm diode laser in conjunction with indocyanine green (ICG)-doped albumin protein solders to repair bovine aorta specimens. Liquid and solid protein solders prepared from 25% and 60% bovine serum albumin (BSA), respectively, were compared. The efficacy of temperature feedback control in enhancing the soldering process was also investigated. Increasing the BSA concentration from 25% to 60% greatly increased the tensile strength of the repairs. A reduction in dye concentration from 2.5 mg ml(-1) to 0.25 mg ml(-1) was also found to result in an increase in tensile strength. Increasing the laser irradiance and thus surface temperature resulted in an increased severity of histological injury. Thermal denaturation of tissue collagen and necrosis of the intimal layer smooth muscle cells increased laterally and in depth with higher temperatures. The strongest repairs were produced with an irradiance of 6.4 W cm(-2) using a solid protein solder composed of 60% BSA and 0.25 mg ml(-1) ICG. Using this combination of laser and solder parameters, surface temperatures were observed to reach 85+/-5 degrees C with a maximum temperature difference through the 150 microm thick solder strips of about 15 degrees C. Histological examination of the repairs formed using these parameters showed negligible evidence of collateral thermal damage to the underlying tissue. Scanning electron microscopy suggested albumin intertwining within the tissue collagen matrix and subsequent fusion with the collagen as the mechanism for laser tissue soldering. The laser tissue soldering technique is shown to be an effective method for producing repairs with improved tensile strength and minimal collateral thermal damage over conventional laser tissue welding techniques.