Lessons learned from the development of a hidradenitis suppurativa xenograft mouse model.

Hidradenitis suppurativa (HS) is a chronic inflammatory skin disease originating from the pilosebaceous unit, in which patients develop painful abscesses, sinus tracts, nodules and scarring, typically in intertriginous areas. Major gaps in our understanding of HS exist, and these may be partially due to the lack of an animal model for experimental studies. We developed an HS xenograft mouse model using human HS lesions grafted onto immunocompromised mice. Although the model had its limitations, several informative lessons were learned, which may contribute to future attempts at an HS animal model.

Specimen Collection for Translational Studies in Hidradenitis Suppurativa.

Hidradenitis suppurativa (HS) is a chronic inflammatory disorder characterized by painful nodules, sinus tracts, and scars occurring predominantly in intertriginous regions. The prevalence of HS is currently 0.053-4%, with a predominance in African-American women and has been linked to low socioeconomic status. The majority of the reported literature is  retrospective, population based, epidemiologic studies. In this regard, there is a need to establish a repository of biospecimens, which represent appropriate gender and racial demographics amongst HS patients. These efforts will diminish knowledge gaps in understanding the disease pathophysiology. Hence, we sought to outline a step-by-step protocol detailing how we established our HS biobank to facilitate the formation of other HS tissue banks. Equipping researchers with carefully detailed processes for collection of HS specimens would accelerate the accumulation of well-organized human biological material. Over time, the scientific community will have access to a broad range of HS tissue biospecimens, ultimately leading to more rigorous basic and translational research. Moreover, an improved understanding of the pathophysiology is necessary for the discovery of novel therapies for this debilitating disease. We aim to provide high impact translational research methodology for cutaneous biology research and foster multidisciplinary collaboration and advancement of our understanding of cutaneous diseases.

Satisfaction, function and repair integrity after arthroscopic versus mini-open rotator cuff repair.

AIMS: Advances in arthroscopic techniques for rotator cuff repair have made the mini-open approach less popular. However, the mini-open approach remains an important technique for repair for many surgeons. The aims of this study were to compare the integrity of the repair, the function of the shoulder and satisfaction post-operatively using these two techniques in patients aged > 50 years. PATIENTS AND METHODS: We identified 22 patients treated with mini-open and 128 patients treated with arthroscopic rotator cuff repair of July 2007 and June 2011. The mean follow-up was two years (1 to 5). Outcome was assessed using the American Shoulder and Elbow Surgeons (ASES) and Simple Shoulder Test (SST) scores, and satisfaction. The integrity of the repair was assessed using ultrasonography. A power analysis ensured sufficient enrolment. RESULTS: There was no statistically significant difference between the age, function, satisfaction, or pain scores (p > 0.05) of the two groups. The integrity of the repair and the mean SST scores were significantly better in the mini-open group (91% of mini-open repairs were intact versus 60% of arthroscopic repairs, p = 0.023; mean SST score 10.9 (standard deviation (sd) 1.3) in the mini-open group; 8.9 (sd 3.5) in arthroscopic group; p = 0.003). The ASES scores were also higher in the mini-open group (mean ASES score 91.0 (sd 10.5) in mini-open group; mean 82.70 (sd 19.8) in the arthroscopic group; p = 0.048). CONCLUSION: The integrity of the repair and function of the shoulder were better after a mini-open repair than after arthroscopic repair of a rotator cuff tear in these patients. The functional difference did not translate into a difference in satisfaction. Mini-open rotator cuff repair remains a useful technique despite advances in arthroscopy. Cite this article: Bone Joint J 2017;99-B:245-9.

Oncogenic Ras/Src cooperativity in pancreatic neoplasia.

Pancreas cancer is one of the most lethal malignancies and is characterized by activating mutations of Kras, present in 95% of patients. More than 60% of pancreatic cancers also display increased c-Src activity, which is associated with poor prognosis. Although loss of tumor suppressor function (for example, p16, p53, Smad4) combined with oncogenic Kras signaling has been shown to accelerate pancreatic duct carcinogenesis, it is unclear whether elevated Src activity contributes to Kras-dependent tumorigenesis or is simply a biomarker of disease progression. Here, we demonstrate that in the context of oncogenic Kras, activation of c-Src through deletion of C-terminal Src kinase (CSK) results in the development of invasive pancreatic ductal adenocarcinoma (PDA) by 5-8 weeks. In contrast, deletion of CSK alone fails to induce neoplasia, while oncogenic Kras expression yields PDA at low frequency after a latency of 12 months. Analysis of cell lines derived from Ras/Src-induced PDA's indicates that oncogenic Ras/Src cooperativity may lead to genomic instability, yet Ras/Src-driven tumor cells remain dependent on Src signaling and as such, Src inhibition suppresses growth of Ras/Src-driven tumors. These findings demonstrate that oncogenic Ras/Src cooperate to accelerate PDA onset and support further studies of Src-directed therapies in pancreatic cancer.

Determination of ankle muscle power in normal gait using an EMG-to-force processing approach.

The purpose of this study was to determine the contribution of individual ankle muscles to the net ankle power and to examine each muscle's role in propulsion or support of the body during normal, self-selected-speed walking. An EMG-to-force processing (EFP) model was developed which scaled muscle tendon unit force output to gait EMG, with that muscle's power output being the product of muscle force and contraction velocity. Net EFP power was determined by summing individual ankle muscle power. Net ankle power was also calculated for these subjects via inverse dynamics. Closeness of fit of the power curves of the two methods was used to validate the model. The curves were highly correlated (r(2)=.91), thus the model was deconstructed to analyze the power contribution and role of each ankle muscle during normal gait. Key findings were that the plantar flexors control tibial rotation in single support, and act to propel the entire limb into swing phase. The dorsiflexors provide positive power for swing phase foot clearance, negative power to control early stance phase foot placement, and a second positive power burst to actively advance the tibia in the transition from double to single support. Co-contraction of agonists and antagonists was limited to only a small percentage of the gait cycle.

Influence of microparticle formulation on immunogenicity of SYI, a synthetic peptide derived from Streptococcus mutans GbpB.

Subcutaneous immunization with SYI, a peptide construct based on Streptococcus mutans glucan binding protein B (GbpB) residues 113-132, significantly reduces experimental dental caries. Since mucosal immunization may be preferred for human vaccine applications, the present objective was to determine what formulation of SYI combined with polylactide-coglycolide microparticles could give rise to significant levels of salivary IgA antibody reactive with the native GbpB protein. A comparison of the SYI construct, loaded into or mixed with polylactide-coglycolide revealed the SYI-loaded microparticles to induce significant and sustainable levels of salivary and nasal wash IgA antibody to the peptide and the native protein. SYI mixed with unloaded microparticles was less effective in mucosal antibody response induction. These studies indicate that mucosal immunization with the SYI construct can induce salivary IgA antibody to a pathogenesis-associated component of S. mutans if delivered within polylactide-coglycolide microparticles, suggesting that this approach could successfully induce protective salivary immunity to dental caries caused by S. mutans.

Remote glucosyltransferase-microparticle vaccine delivery induces protective immunity in the oral cavity.

Intranasally administered dental caries vaccines show significant promise for human application. Alternate mucosal routes may be required, however, to induce caries-protective salivary IgA antibody in children with respiratory diseases. Since rectal mucosa contains inductive lymphoid tissue, we hypothesized that the rectal route could be used to induce salivary immunity to mutans streptococcal glucosyltransferase (GTF), resulting in protective immunity to experimental dental caries. We first explored the ability of glucosyltransferase, incorporated into polylactide-co-glycolide (PLGA) microparticles (MP), and administered rectally together with mucosal adjuvant, to induce a salivary IgA antibody response. Groups of Sprague-Dawley rats (6/group) were immunized rectally on days 0, 7, 14 and 21 with a) GTF-MP alone, b) GTF-MP with cholera toxin, c) GTF-MP with detoxified mutant Escherichia coli toxin (dLT), or d) sham immunized with PLGA and cholera toxin. An additional group was immunized intranasally with GTF-MP alone. Saliva and nasal washes of all intranasally immunized rats contained IgA antibody to glucosyltransferase on day 28. Salivary IgA antibody was also detected in 7/12 rats rectally immunized with GTF-MP and cholera toxin or dLT, although responses were lower than those obtained by intranasal immunization. Most fecal extracts from rectally delivered GTF-MP plus cholera toxin or dLT rats contained IgA antibody to GTF-MP. Low levels of fecal IgA antibody were detected in 3/6 intranasally immunized rats and 2/6 rats rectally immunized with GTF-MP alone. We then examined the extent to which salivary IgA antibody induced by the rectal route could be protective. At 25, 31 and 38 days of age, two groups of female Sprague-Dawley rats (13/group) were rectally immunized with GTF-MP and cholera toxin or with empty microparticles and cholera toxin (sham group). A third group was intranasally immunized with GTF-MP alone. After demonstrating salivary IgA responses to GTF in most GTF-immunized rats, all animals were infected with streptomycin-resistant Streptococcus sobrinus and placed on diet 2000. After 79 days of infection, total caries on molar surfaces were lower in both rectally (7.9 +/- 1.0) and intranasally (7.1 +/- 0.9; P < 0.0.03) immunized groups compared with the sham-immunized group (11.9 +/- 1.6). Smooth surface caries were significantly lower (P < 0.05) in both rectally and intranasally immunized groups. These results support the interconnectedness of the mucosal immune system and indicate that rectal immunization with GTF-MP, together with adjuvant, or intranasal immunization with GTF-MP alone, can induce protective levels of salivary antibody in rats.

Facilitated intranasal induction of mucosal and systemic immunity to mutans streptococcal glucosyltransferase peptide vaccines.

Synthetic peptide vaccines which are derived from functional domains of Streptococcus mutans glucosyltransferases (GTF) have been shown to induce protective immunity in Sprague-Dawley rats after subcutaneous injection in the salivary gland region. Since mucosal induction of salivary immunity would be preferable in humans, we explored methods to induce mucosal antibody in the rat to the GTF peptide vaccines HDS and HDS-GLU after intranasal administration. Several methods of facilitation of the immune response were studied: the incorporation of peptides in bioadhesive poly(D,L-lactide-coglycolide) (PLGA) microparticles, the use of monoepitopic (HDS) or diepitopic (HDS-GLU) peptide constructs, or the use of mucosal adjuvants. Salivary immunoglobulin A (IgA) responses were not detected after intranasal administration of diepitopic HDS-GLU peptide constructs in alum or after incorporation into PLGA microparticles. However, significant primary and secondary salivary IgA and serum IgG antibody responses to HDS were induced in all rats when cholera holotoxin (CT) or a detoxified mutant Escherichia coli heat-labile enterotoxin (R192G LT) were intranasally administered with HDS peptide constructs in PLGA. Coadministration of LT with HDS resulted in predominantly IgG2a responses in the serum, while coadministration with CT resulted in significant IgG1 and IgG2a responses to HDS. Serum IgG antibody, which was induced to the HDS peptide construct by coadministration with these adjuvants, also bound intact mutans streptococcal GTF in an enzyme-linked immunosorbent assay and inhibited its enzymatic activity. Thus, immune responses which are potentially protective for dental caries can be induced to peptide-based GTF vaccines after mucosal administration if combined with the CT or LT R192G mucosal adjuvant.

Domain organization and flavin adenine dinucleotide-binding determinants in the aerotaxis signal transducer Aer of Escherichia coli.

Aerotactic responses in Escherichia coli are mediated by the membrane transducer Aer, a recently identified member of the superfamily of PAS domain proteins, which includes sensors of light, oxygen, and redox state. Initial studies of Aer suggested that it might use a flavin adenine dinucleotide (FAD) prosthetic group to monitor cellular redox changes. To test this idea, we purified lauryl maltoside-solubilized Aer protein by His-tag affinity chromatography and showed by high performance liquid chromatography, mass spectrometry, and absorbance spectroscopy that it bound FAD noncovalently. Polypeptide fragments spanning the N-terminal 290 residues of Aer, which contains the PAS motif, were able to bind FAD. Fusion of this portion of Aer to the flagellar signaling domain of Tsr, the serine chemoreceptor, yielded a functional aerotaxis transducer, demonstrating that the FAD-binding portion of Aer is sufficient for aerosensing. Aerotaxis-defective missense mutants identified two regions, in addition to the PAS domain, that play roles in FAD binding. Those regions flank a central hydrophobic segment needed to anchor Aer to the cytoplasmic membrane. They might contact the FAD ligand directly or stabilize the FAD-binding pocket. However, their lack of sequence conservation in Aer homologs of other bacteria suggests that they play less direct roles in FAD binding. One or both regions probably also play important roles in transmitting stimulus-induced conformational changes to the C-terminal flagellar signaling domain to trigger aerotactic behavioral responses.

The Rancho EMG analyzer: a computerized system for gait analysis.

This paper describes a computer system which accurately defines the EMG patterns of the lower extremities during gait. Footswitches are used to identify the temporal relationships and determine the phases of the gait cycle. Fine wire electrodes, inserted in the desired muscles of the patient being tested, provide EMG signals for comparison with a normal database. The system is also usable with surface electrodes when an appropriate normal database for surface electrodes is incorporated. Descriptive qualifiers (such as 'premature onset', 'delayed cessation', 'no clinically significant EMG', 'continuous activity' etc.) are used to produce a clinically relevant printed (textual) report. The intensity filtered average (IFA) of the EMG is shown graphically with the representative profile of each stride. The IFAs for all muscles tested can be plotted together (up to six on a page) and the graphic representation of the 'raw' EMG can be produced. The methods of generating the normal database by creating time-adjusted mean profiles (TAMP) are enumerated. The clinical use of the system is discussed. A detailed analysis of 31 of the most recent patient tests for which the system was used provides an indication of its accuracy. For 86% of the 428 muscle tests examined, the EMG analyser was considered to have given the correct result as compared with a visual analysis of the raw EMG record by a trained expert. Recommendations for the use and future improvements of the EMG analyser are made.

A computer algorithm for defining the group electromyographic profile from individual gait profiles.

A computer algorithm was developed to determine the group electromyographic (EMG) profile for the soleus muscle during free speed level walking. Subjects consisted of 50 adults (21 male, 29 female) with no history of musculoskeletal disease. EMG was recorded from the soleus muscle with wire electrodes, and was normalized by maximum muscle test. Two algorithms (time-adjusted mean profile (TAMP) and mean intensity profile (MIP)) were implemented to construct a group profile from identical individual EMG profiles. In addition, a grand ensemble average (GEAV) of the same individual data was performed. A high positive correlation (omega 2 > .995) was found between MIP and GEAV of EMG data. A control group was established based on mean timing and relative intensity of the individual EMG profiles. The MIP and GEAV were shown to have earlier onsets, later cessations, and extended EMG duration in comparison to control values. No significant differences were observed comparing TAMP and mean values for any measure.

Computer algorithms to characterize individual subject EMG profiles during gait.

Three methods of precisely determining onset and cessation times of gait EMG were investigated. Subjects were 24 normal adults and 32 individuals with gait pathologies. Soleus muscle EMG during free speed level walking was obtained with fine wires, and was normalized by manual muscle test (%MMT). Linear envelopes were generated from the rectified, integrated EMG at each percent gait cycle (%GC) of each stride in individual gait trials. Three methods were used to generate EMG profiles for each tested subject. The ensemble average (EAV) was determined for each subject from the mean relative intensity of the linear envelopes. Low relative intensity or short duration EMG was removed from the ensemble average to create the intensity filtered average (IFA). The packet analysis method (PAC) created an EMG profile from the linear envelopes in successive strides whose respective centroid %GC locations were within +/- 15%GC of each other. Control values for onset and cessation times of individual gait trials were calculated after spurious outliers were removed. Mean onset and cessation times across subjects for control values and the experimental methods (EAV, IFA, and PAC) were calculated. Dunnett's test (p less than .05) was performed to compare control and experimental groups in patient and normal trials. EAV differed from control values for onsets (p less than .01), cessations (p less than .01), and durations (p less than .01) in both normal and patient trials. IFA and PAC had no significant differences from control value means. IFA was selected for clinical use as automatic analysis could be performed on all trials and a minimum number of decision rules were needed.

Ectopic decidual reaction of the uterine cervix: frequency and cytologic presentation.

The histologic and cytologic manifestation of cervical decidual reaction in pregnancy was investigated. Histologically, cervical decidua was found in 30.8% of 191 uteri removed during pregnancy. Characteristic features were focal occurrence, subepithelial location and frequent disruption of the overlying epithelium. Cytologically, decidual cells were found in 34% of patients with histologically confirmed cervical decidual reaction. The number of decidual cells per smear ranged from 11 to 208, with a mean of 105. The majority of cells occurred in aggregates and stained basophilic or amphophilic. Outstanding diagnostic features of decidual cells in smears were finely granular chromatin pattern in 96%, prominent nucleoli in over 80% and marked nuclear enlargement. Cytometrically, decidual cells showed a wide variation in size. The nuclear area ranged from 31 sq micrometers to 320 sq micrometers, with a mean of 111 sq micrometers. In the differential diagnosis, tissue repair, dysplasia, carcinoma in situ and endocervical adenocarcinoma must be considered.

Functional evaluation of the pes anserinus transfer by electromyography and gait analysis.

The effectiveness of the pes anserinus transfer, proposed by Slocum and Larson to control anteromedial rotatory instability of the knee, depends on the pes anserinus muscles contracting at times when the instability threatens the patient's function. To examine the activity of the transferred pes anserinus muscles, quantitated dynamic electromyography, isometric testing of muscle strength, and gait-analysis techniques were employed. Seven normal subjects and nine patients who had undergone pes anserinus transfer for anteromedial rotatory instability were tested. In contrast to the normal subjects, who demonstrated pes anserinus muscle activity primarily during swing phase, the patients displayed conspicuous activity of the semitendinosus, gracilis, and sartorius tendons during stance phase. Quantitated electromyography indicated that the pes anserinus muscles worked harder in the patients than in the normal subjects, and more effort was required from the muscles of patients with poor results than from those of patients with good results. All of the patients also demonstrated reduction of single-limb support time, as well as a shorter stride length and reduced gait velocity. We concluded that the pes anserinus transfer is kinetically sound. Since instability of the knee is a functional defect that is present during stance, the finding of marked activity of the pes anserinus muscles during stance phase in the patients but not in the normal subjects suggests that these muscles were being used to control the instability.