Increased actual sling wear is associated with better early patient-reported and image-based outcomes after shoulder surgery.

BACKGROUND: Prior studies have failed to show differences in functional outcomes for patient-reported sling use after rotator cuff repair. Temperature-sensing devices are used to more accurately measure brace adherence. The purposes of this study were to quantify actual sling adherence and its predictors and to establish whether increased sling adherence is associated with improved functional and image-based outcomes. METHODS: We performed a prospective cohort study of 65 patients undergoing shoulder surgery requiring ≥4 weeks of postoperative sling use. Temperature-sensing devices were implanted in the slings to monitor sling adherence. Patient-reported sling adherence was determined from a questionnaire. Patients were considered 80% adherent if they wore the sling 16 h/d (112 h/week) when 20 h/d was prescribed. The primary outcomes were patient-reported and actual sling adherence, patient-reported outcomes (American Shoulder and Elbow Surgeons score and visual analog scale pain score) within 12 months postoperatively, and image-based failure based on ultrasound or radiography at 6 weeks and 1 year postoperatively. RESULTS: Patient-reported sling adherence was highly sensitive (82.8%), was poorly specific (28.6%), had low accuracy (53.1%), and was weakly correlated with actual sling adherence (r = 0.32, P = .009). On multivariable logistic regression analysis, male patients were 91% less likely than female patients to be adherent with sling use (odds ratio, 0.09; 95% confidence interval [CI], 0.02-0.42; P = .002). Additionally, obese and morbidly obese patients were 88% (95% CI, 0.02-0.84; P = .033) and 98% (95% CI, 0.002-0.27; P = .003), respectively, less likely than non-obese patients to adhere to sling wear postoperatively. After we controlled for surgical procedure, visual analog scale pain scores were significantly better at 6 weeks (ß = -1.47; 95% CI, -2.88 to -0.05; P = .04) and 3 months (ß = -1.68; 95% CI, -3.28 to -0.08; P = .04) if patients adhered to sling wear. A receiver operating characteristic curve showed that 13.6 hours and 15.4 hours of daily sling wear optimized image-based outcomes at 6 weeks (failure rate, 0% vs. 16%; P = .01) and 1 year (failure rate, 3% vs. 28%; P = .008) postoperatively, respectively. CONCLUSION: The results of this study demonstrate that patient-reported sling adherence is unreliable, adherence can be predicted by female sex and lower body mass index, and increased sling adherence is associated with improved early pain scores and image-based outcomes. These data can help inform future studies using postoperative sling protocols as patient-reported sling adherence is not an accurate method to assess sling use.

The Effect of Patients' Understanding of Sling Necessity and Home Assistance on Sling Wear After Shoulder Surgery.

BACKGROUND: Perioperative education and socioeconomic factors influence patient behavior. Recent evidence has suggested that sling compliance is associated with improved outcomes after shoulder surgery; it is important to investigate factors that influence sling compliance. PURPOSE: To determine the associations between postoperative sling wear and patients' understanding of sling necessity, postoperative home assistance, and social deprivation. STUDY DESIGN: Cohort study; Level of evidence, 2. METHODS: A total of 66 patients were prospectively enrolled from 2018 to 2020 if they were ≥18 years of age and undergoing shoulder surgery requiring a sling for at least 1 month postoperatively. Sling wear was measured using a temperature-sensing device. At 6 weeks postoperatively, patients' understanding for sling necessity was determined by their response to a question on the Medical Adherence Measure questionnaire, "Why did you have to wear a shoulder sling?" The Patient Understanding Grading Scale (PUGS) was developed to quantify patient responses. PUGS was graded 1 to 3, with grade 1 corresponding to the least technical knowledge. Patient characteristics, social deprivation (Area Deprivation Index [ADI]), and home assistance were additionally analyzed. RESULTS: There were no significant differences in baseline characteristics between patients when stratified by PUGS grade. Multivariable linear regression analysis for total hours of sling wear per week showed that patients with PUGS grade 2 (ß, 48.2 hours; P = .007) and grade 3 (ß, 59.5 hours; P = .003) wore their slings significantly more than grade 1 patients. Patients with home assistance had significantly greater day hours (73.5 ± 33.0 vs 44.0 ± 24.5 hours; P = .037) of sling wear per week, but there was no difference in night sling hours. Patients older than 60 years wore their slings significantly more, while men and those with a higher body mass index (BMI) wore their slings significantly less. ADI was not significantly associated with sling wear. CONCLUSION: This study demonstrates that patients with greater understanding for sling necessity, those with home assistance, and patients >60 years have greater sling wear, while male patients and those with a higher BMI have lower sling compliance. ADI was not a significant contributor.

Fibroadipogenic progenitor cell response peaks prior to progressive fatty infiltration after rotator cuff tendon tear.

Fibroadipogenic progenitor (FAP) cells are implicated as a major source of fatty infiltration (FI) in murine rotator cuff (RC) injury, but FAP cell response after RC tear in a rabbit model is unknown. This study determined whether changes in FAP cell count after an RC tear predate muscle degeneration in a clinically relevant rabbit model. We hypothesized increases in FAP cell count correlate temporally with RC degeneration. New Zealand white rabbits (n = 26) were evaluated at 1, 2, 4, and 6 weeks after unilateral full-thickness tenotomy of supraspinatus and infraspinatus tendons. FI area and adipocyte size were histologically analyzed, muscle density was measured by computerized tomography, and quantification of FAP cells was measured by flow cytometry and immunohistochemistry. The percentage of intrafascicular adipocyte area increased over time in supraspinatus muscle samples (p = 0.03), significantly between 1- and 6-week samples (p = 0.04). There were no differences in perifascicular adipocyte area percentages between time points. Peak increase in FAP cell count occurred at 1-week (p = 0.03), with a decrease in the following weeks. There was a negative correlation between supraspinatus adipocyte area and FAP cell count (p < 0.05). On computed tomography (CT) scan, maximal decrease in muscle density was observed in the 4th to 6th weeks. In summary, FAP cell response occurred early after tenotomy and did not correlate temporally with increases in FI. This suggests that FAP cell response may predate degenerative changes, and early targeting of FAPs before adipocyte maturation could blunt FI after RC tear.

Disparate bone anabolic cues activate bone formation by regulating the rapid lysosomal degradation of sclerostin protein.

The downregulation of sclerostin in osteocytes mediates bone formation in response to mechanical cues and parathyroid hormone (PTH). To date, the regulation of sclerostin has been attributed exclusively to the transcriptional downregulation of the Sost gene hours after stimulation. Using mouse models and rodent cell lines, we describe the rapid, minute-scale post-translational degradation of sclerostin protein by the lysosome following mechanical load and PTH. We present a model, integrating both new and established mechanically and hormonally activated effectors into the regulated degradation of sclerostin by lysosomes. Using a mouse forelimb mechanical loading model, we find transient inhibition of lysosomal degradation or the upstream mechano-signaling pathway controlling sclerostin abundance impairs subsequent load-induced bone formation by preventing sclerostin degradation. We also link dysfunctional lysosomes to aberrant sclerostin regulation using human Gaucher disease iPSCs. These results reveal how bone anabolic cues post-translationally regulate sclerostin abundance in osteocytes to regulate bone formation.

ProxiMAX randomization: a new technology for non-degenerate saturation mutagenesis of contiguous codons.

Back in 2003, we published 'MAX' randomization, a process of non-degenerate saturation mutagenesis using exactly 20 codons (one for each amino acid) or else any required subset of those 20 codons. 'MAX' randomization saturates codons located in isolated positions within a protein, as might be required in enzyme engineering, or else on one face of an α-helix, as in zinc-finger engineering. Since that time, we have been asked for an equivalent process that can saturate multiple contiguous codons in a non-degenerate manner. We have now developed 'ProxiMAX' randomization, which does just that: generating DNA cassettes for saturation mutagenesis without degeneracy or bias. Offering an alternative to trinucleotide phosphoramidite chemistry, ProxiMAX randomization uses nothing more sophisticated than unmodified oligonucleotides and standard molecular biology reagents. Thus it requires no specialized chemistry, reagents or equipment, and simply relies on a process of saturation cycling comprising ligation, amplification and digestion for each cycle. The process can encode both unbiased representation of selected amino acids or else encode them in predefined ratios. Each saturated position can be defined independently of the others. We demonstrate accurate saturation of up to 11 contiguous codons. As such, ProxiMAX randomization is particularly relevant to antibody engineering.

Sociocultural practices in Ethiopia: association with onset and persistence of postnatal common mental disorders.

BACKGROUND: Traditional perinatal practices may protect against postnatal common mental disorders (CMD) in non-Western societies. AIMS: To evaluate the association between perinatal practices and postnatal CMD in rural Ethiopia. METHOD: A population-based sample of 1065 women was followed up from pregnancy until 2 months postpartum. Qualitative investigation informed the development of scales measuring attitudes towards and adherence to perinatal practices. Postnatal CMD was measured using the Self-Reporting Questionnaire. RESULTS: Endorsement of sociocultural perinatal practices was associated with lower odds of antenatal CMD persisting into the postnatal period (adjusted odds ratio (OR) = 0.66, 95% CI 0.45-0.95). Women who endorsed protective and celebratory perinatal practices but were unable to complete them had increased odds of incident (adjusted OR = 7.26, 95% CI 1.38-38.04) and persistent postnatal CMD (adjusted OR = 2.16, 95% CI 1.11-4.23) respectively. CONCLUSIONS: There is evidence for an independent role of sociocultural practices in maintaining perinatal mental health in this Ethiopian community.

The effect of maternal common mental disorders on infant undernutrition in Butajira, Ethiopia: the P-MaMiE study.

BACKGROUND: Although maternal common mental disorder (CMD) appears to be a risk factor for infant undernutrition in South Asian countries, the position in sub-Saharan Africa (SSA) is unclear METHODS: A population-based cohort of 1065 women, in the third trimester of pregnancy, was identified from the demographic surveillance site (DSS) in Butajira, to investigate the effect of maternal CMD on infant undernutrition in a predominantly rural Ethiopian population. Participants were interviewed at recruitment and at two months post-partum. Maternal CMD was measured using the locally validated Self-Reported Questionnaire (score of > or = six indicating high levels of CMD). Infant anthropometry was recorded at six and twelve months of age. RESULT: The prevalence of CMD was 12% during pregnancy and 5% at the two month postnatal time-point. In bivariate analysis antenatal CMD which had resolved after delivery predicted underweight at twelve months (OR = 1.71; 95% CI: 1.05, 2.50). There were no other statistically significant differences in the prevalence of underweight or stunted infants in mothers with high levels of CMD compared to those with low levels. The associations between CMD and infant nutritional status were not significant after adjusting for pre-specified potential confounders. CONCLUSION: Our negative finding adds to the inconsistent picture emerging from SSA. The association between CMD and infant undernutrition might be modified by study methodology as well as degree of shared parenting among family members, making it difficult to extrapolate across low- and middle-income countries.

Optical fibre-based detection of DNA hybridization.

A dual-peak LPFG (long-period fibre grating), inscribed in an optical fibre, has been employed to sense DNA hybridization in real time, over a 1 h period. One strand of the DNA was immobilized on the fibre, while the other was free in solution. After hybridization, the fibre was stripped and repeated detection of hybridization was achieved, so demonstrating reusability of the device. Neither strand of DNA was fluorescently or otherwise labelled. The present paper will provide an overview of our early-stage experimental data and methodology, examine the potential of fibre gratings for use as biosensors to monitor both nucleic acid and other biomolecular interactions and then give a summary of the theory and fabrication of fibre gratings from a biological standpoint. Finally, the potential of improving signal strength and possible future directions of fibre grating biosensors will be addressed.

Impact of antenatal common mental disorders upon perinatal outcomes in Ethiopia: the P-MaMiE population-based cohort study.

OBJECTIVES: To examine the impact of antenatal psychosocial stressors, including maternal common mental disorders (CMD), upon low birth weight, stillbirth and neonatal mortality, and other perinatal outcomes in rural Ethiopia. METHODS: A population-based sample of 1065 pregnant women was assessed for symptoms of antenatal CMD (Self-Reporting Questionnaire-20: SRQ-20), stressful life events during pregnancy (List of Threatening Experiences: LTE) and worry about the forthcoming delivery. In a sub-sample of 654 women from six rural sub-districts, neonatal birth weight was measured on 521 (79.7%) singleton babies within 48 h of delivery. Information about other perinatal outcomes was obtained shortly after birth from the mother's verbal report and via the Demographic Surveillance System. RESULTS: After adjusting for potential confounders, none of the psychosocial stressors were associated with lower mean birth weight, stillbirth or neonatal mortality. Increasing levels of antenatal CMD symptoms were associated both with prolonged labour (>24 h) (SRQ 1-5: RR 1.4; 95% CI 1.0-1.9, SRQ >or= 6: RR 1.6; 95% CI 1.0-2.6) and delaying initiation of breast-feeding more than eight hours (SRQ 1-5: RR 1.4; 95% CI 0.8 to 2.4, SRQ >or= 6: RR 2.8; 95% CI 1.3-6.1). Worry about delivery was also associated with labour longer than 24 h (RR 1.5; 95% CI 1.0-2.1). CONCLUSIONS: This study provides preliminary evidence of important public health consequences of poor maternal mental health in low-income countries but does not replicate the strong association with low birth weight found in South Asia.

Development of human cloned blastocysts following somatic cell nuclear transfer with adult fibroblasts.

Nuclear transfer stem cells hold considerable promise in the field of regenerative medicine and cell-based drug discovery. In this study, a total of 29 oocytes were obtained from three young (20-24 years old) reproductive egg donors who had been successful in previous cycles. These oocytes, deemed by intended parents to be in excess of their reproductive needs, were donated for research without financial compensation by both the egg donor and intended parents after receiving informed consent. All intended parents successfully achieved ongoing pregnancies with the oocytes retained for reproductive purposes. Mature oocytes, obtained within 2 hours following transvaginal aspiration, were enucleated using one of two methods, extrusion or aspiration, after 45 minutes of incubation in cytochalasin B. Rates of oocyte lysis or degeneration did not differ between the two methods. Somatic cell nuclear transfer (SCNT) embryos were constructed using two established adult male fibroblast lines of normal karyotype. High rates of pronuclear formation (66%), early cleavage (47%), and blastocyst (23%) development were observed following incubation in standard in vitro fertilization culture media. One cloned blastocyst was confirmed by DNA and mitochondrial DNA fingerprinting analyses, and DNA fingerprinting of two other cloned blastocysts indicated that they were also generated by SCNT. Blastocysts were also obtained from a limited number of parthenogenetically activated oocytes. This study demonstrates, for the first time, that SCNT can produce human blastocyst-stage embryos using nuclei obtained from differentiated adult cells and provides new information on methods that may be needed for a higher level of efficiency for human nuclear transfer.

Detecting perinatal common mental disorders in Ethiopia: validation of the self-reporting questionnaire and Edinburgh Postnatal Depression Scale.

BACKGROUND: The cultural validity of instruments to detect perinatal common mental disorders (CMD) in rural, community settings has been little-investigated in developing countries. METHODS: Semantic, content, technical, criterion and construct validity of the Edinburgh Postnatal Depression Scale (EPDS) and Self-Reporting Questionnaire (SRQ) were evaluated in perinatal women in rural Ethiopia. Gold-standard measure of CMD was psychiatric assessment using the Comprehensive Psychopathological Rating Scale (CPRS). Community-based, convenience sampling was used. An initial validation study (n=101) evaluated both EPDS and SRQ. Subsequent validation was of SRQ alone (n=119). RESULTS: EPDS exhibited poor validity; area under the receiver operating characteristic (AUROC) curve of 0.62 (95%CI 0.49 to 0.76). SRQ-20 showed better validity as a dimensional scale, with AUROC of 0.82 (95%CI 0.68 to 0.96) and 0.70 (95%CI 0.57 to 0.83) in the two studies. The utility of SRQ in detecting 'cases' of CMD was not established, with differing estimates of optimal cut-off score: three and above in Study 1 (sensitivity 85.7%, specificity 75.6%); seven and above in Study 2 (sensitivity 68.4%, specificity 62%). High convergent validity of SRQ as a dimensional measure was demonstrated in a community survey of 1065 pregnant women. LIMITATIONS: Estimation of optimal cut-off scores and validity coefficients for detecting CMD was limited by sample size. CONCLUSIONS: EPDS demonstrated limited clinical utility as a screen for perinatal CMD in this rural, low-income setting. The SRQ-20 was superior to EPDS across all domains for evaluating cultural equivalence and showed validity as a dimensional measure of perinatal CMD.

Real-time detection of DNA interactions with long-period fiber-grating-based biosensor.

Using an optical biosensor based on a dual-peak long-period fiber grating, we have demonstrated the detection of interactions between biomolecules in real time. Silanization of the grating surface was successfully realized for the covalent immobilization of probe DNA, which was subsequently hybridized with the complementary target DNA sequence. It is interesting to note that the DNA biosensor was reusable after being stripped off the hybridized target DNA from the grating surface, demonstrating a function of multiple usability.

Discovery of active proteins directly from combinatorial randomized protein libraries without display, purification or sequencing: identification of novel zinc finger proteins.

We have successfully linked protein library screening directly with the identification of active proteins, without the need for individual purification, display technologies or physical linkage between the protein and its encoding sequence. By using 'MAX' randomization we have rapidly constructed 60 overlapping gene libraries that encode zinc finger proteins, randomized variously at the three principal DNA-contacting residues. Expression and screening of the libraries against five possible target DNA sequences generated data points covering a potential 40,000 individual interactions. Comparative analysis of the resulting data enabled direct identification of active proteins. Accuracy of this library analysis methodology was confirmed by both in vitro and in vivo analyses of identified proteins to yield novel zinc finger proteins that bind to their target sequences with high affinity, as indicated by low nanomolar apparent dissociation constants.

Fluorescent microplate-based analysis of protein-DNA interactions. II: Immobilized DNA.

A simple protein-DNA interaction analysis has been developed using both a high-affinity/high-specificity zinc finger protein and a low-specificity zinc finger protein with nonspecific DNA binding capability. The latter protein is designed to mimic background binding by proteins generated in randomized or shuffled gene libraries. In essence, DNA is immobilized onto the surface of microplate wells via streptavidin capture, and green fluorescent protein (GFP)-labeled protein is added in solution as part of a crude cell lysate or protein mixture. After incubation and washing, bound protein is detected in a standard microplate reader. The minimum sensitivity of the assay is approximately 0.4 nM protein. The assay format is ideally suited to investigate the interactions of DNA binding proteins from within crude cell extracts and/or mixtures of proteins that may be encountered in protein libraries generated by codon randomization or gene shuffling.

Messenger RNA expression profiling of genes involved in epidermal growth factor receptor signalling in human cancer cells treated with scanning array-designed antisense oligonucleotides.

Scanning oligodeoxynucleotide (ODN) arrays appear promising in vitro tools for the prediction of effective antisense reagents but their usefulness has not yet been reported in mammalian systems. In this study, we have evaluated the use of scanning ODN arrays to predict efficacious antisense ODNs targeting the human epidermal growth factor receptor (EGFR) mRNA in a human epidermoid cancer cell line and in primary human glioma cells. Hybridisation accessibility profile of the first 120nt in the coding region of the human EGFR mRNA was determined by hybridising a radiolabelled EGFR transcript to a scanning array of 2684 antisense sequences ranging from monomers to 27-mers. Two ODNs, AS1 and AS2, complementary to accessible sequences within the EGFR mRNA, were designed and their ability to hybridise to EGFR mRNA was further confirmed by in vitro RNase H-mediated cleavage assays. Phosphorothioate-modified 21-mer AS1 and AS2 ODNs inhibited the growth of an established human A431 cancer cell line as well as primary glioma cells from human subjects when delivered as cationic lipoplexes. In contrast, scrambled controls and AS3-an antisense ODN complementary to an inaccessible site in EGFR mRNA-were inactive. Western blots showed that AS1 ODN exhibited a dose-dependent inhibition of EGFR protein expression in A431 cells in the nanomolar range. Microarray-based gene expression profiling studies of A431 cells treated with the 21-mer phosphorothioate AS1 ODN demonstrated successful inhibition of downstream signalling molecules further confirming the effective inhibition of EGFR expression in human cancer cells by antisense ODNs designed by scanning ODN array technology.