Health care providers' weight management practices for adolescent obesity and alignment with clinical practice guidelines: a multi-centre, qualitative study.

BACKGROUND: Clinical practice guidelines (CPGs) include evidence-based recommendations for managing obesity in adolescents. However, information on how health care providers (HCPs) implement these recommendations in day-to-day practice is limited. Our objectives were to explore how HCPs deliver weight management health services to adolescents with obesity and describe the extent to which their reported practices align with recent CPGs for managing pediatric obesity. METHODS: From July 2017 to January 2018, we conducted a qualitative study that used purposeful sampling to recruit HCPs with experience in adolescent weight management from multidisciplinary, pediatric weight management clinics in Edmonton and Ottawa, Canada. Data were collected using audio-recorded focus groups (4-6 participants/group; 60-90 min in length). We applied inductive, semantic thematic analysis and the congruent methodological approach to analyze our data, which included transcripts, field notes, and memos. Qualitative data were compared to recent CPGs for pediatric obesity that were published by the Endocrine Society in 2017. Of the 12 obesity 'treatment-related' recommendations, four were directly relevant to the current study. RESULTS: Data were collected through three focus groups with 16 HCPs (n = 10 Edmonton; n = 6 Ottawa; 94% female; 100% Caucasian), including dietitians, exercise specialists, nurses, pediatricians, psychologists, and social workers. We identified three main themes that we later compared with CPG recommendations, including: (i) discuss realistic expectations regarding weight management (e.g., shift focus from weight to health; explore family cohesiveness; foster delayed vs instant gratification), (ii) personalize weight management (e.g., address personal barriers to change; consider developmental readiness), and (iii) exhibit non-biased attitudes and practices (e.g., de-emphasize individual causes of obesity; avoid making assumptions about lifestyle behaviors based on weight). Based on these qualitative findings, HCPs applied all four CPG recommendations in their practices. CONCLUSIONS: HCPs provided practical insights into what and how they delivered weight management for adolescents, which included operationalizing relevant CPG recommendations in their practices.

End-user perspectives to inform policy and program decisions: a qualitative and quantitative content analysis of lifestyle treatment recommendations by adolescents with obesity.

BACKGROUND: Lifestyle modifications represent the first line of treatment in obesity management; however, many adolescents with obesity do not meet lifestyle recommendations. Given that adolescents are rarely consulted during health policy development and in the design of lifestyle interventions, their first-hand experiences, preferences, and priorities may not be represented. Accordingly, our purpose was to explore adolescents' lifestyle treatment recommendations to inform policy and program decisions. METHODS: Conducted from July 2017 to January 2018, this study adhered to a qualitative, crosslanguage, patient-oriented design. We recruited 19 13-17-year-old adolescents (body mass index [BMI] ≥85th percentile) seeking multidisciplinary treatment for obesity in geographically and culturally diverse regions of Canada. Adolescents participated in one-on-one, in-person, semi-structured interviews in English or French. Interviews were audio-recorded, transcribed verbatim, managed using NVivo 11, and analyzed using quantitative and qualitative content analysis by two independent researchers. RESULTS: Adolescents' recommendations were organized into five categories, each of which denotes health as a collective responsibility: (i) establish parental support within limits, (ii) improve accessibility and availability of 'healthy foods', (iii) limit deceptive practices in food marketing, (iv) improve accessibility and availability of varied physical activity opportunities, and (v) delay school start times. Respect for individual autonomy and decision-making capacity were identified as particularly important, however these were confronted with adolescents' partial knowledge on nutrition and food literacy. CONCLUSIONS: Adolescents' recommendations highlighted multi-level, multi-component factors that influenced their ability to lead healthy lifestyles. Uptake of these recommendations by policy-makers and program developers may be of added value for lifestyle treatment targeting adolescents with obesity.

"Fat is really a four-letter word": Exploring weight-related communication best practices in children with and without disabilities and their caregivers.

BACKGROUND: Health care professionals play a critical role in preventing and managing childhood obesity, but the American Academy of Pediatrics recently stressed the importance of using sensitive and nonstigmatizing language when discussing weight with children and families. Although barriers to weight-related discussions are well known, there are few evidence-based recommendations around communication best practices. Disability populations in particular have previously been excluded from work in this area. The objectives were to present the findings of a recent scoping review to children with and without disabilities and their caregivers for their reactions; and to explore the experiences and perceptions of the children and their caregivers regarding weight-related communication best practices. METHODS: Focus group and individual interviews were conducted with 7-18-year olds with and without disabilities and their caregivers. The interview guide was created using findings from a recent scoping review of weight-related communication best practices. Inductive thematic analysis was employed. RESULTS: Eighteen children (9 boys; 7 children with disabilities) and 21 caregivers (17 mothers, 1 step-father, 3 other caregivers) participated in 8 focus group and 7 individual interviews. Preferred communication strategies were similar across those with and without disabilities, although caregivers of children with autism spectrum disorder endorsed more concrete approaches. Discussions emphasizing growth and health were preferred over weight and size. Strengths-based, solution-focused approaches for weight conversations were endorsed, although had not been widely experienced. CONCLUSION: Perceptions of weight-related communication were similar across stakeholder groups, regardless of children's disability or weight status. Participants generally agreed with the scoping review recommendations, suggesting that they apply broadly across different settings and populations; however, tailoring them to specific circumstances is critical. Empirical evaluations are still required to examine the influence of weight-related communication on clinically important outcomes, including behaviour change and family engagement in care.

Why do families enrol in paediatric weight management? A parental perspective of reasons and facilitators.

BACKGROUND: Few children with obesity who are referred for weight management end up enroled in treatment. Factors enabling enrolment are poorly understood. Our purpose was to explore reasons for and facilitators of enrolment in paediatric weight management from the parental perspective. METHODS: Semi-structured interviews were conducted with parents of 10- to 17-year-olds who were referred to one of four Canadian weight management clinics and enroled in treatment. Interviews were audio-recorded and transcribed verbatim. Manifest/inductive content analysis was used to analyse the data, which included the frequency with which parents referred to reasons for and facilitators of enrolment. RESULTS: In total, 65 parents were interviewed. Most had a child with a BMI ≥95th percentile (n = 59; 91%), were mothers (n = 55; 85%) and had completed some post-secondary education (n = 43; 66%). Reasons for enrolment were related to concerns about the child, recommended care and expected benefits. Most common reasons included weight concern, weight loss expectation, lifestyle improvement, health concern and need for external support. Facilitators concerned the referral initiator, treatment motivation and barrier control. Most common facilitators included the absence of major barriers, parental control over the decision to enrol, referring physicians stressing the need for specialized care and parents' ability to overcome enrolment challenges. CONCLUSIONS: Healthcare providers might optimize enrolment in paediatric weight management by being proactive in referring families, discussing the advantages of the recommended care to meet treatment expectations and providing support to overcome enrolment barriers.

Vigorous physical activity and longitudinal associations with cardiometabolic risk factors in youth.

OBJECTIVE: To examine the longitudinal associations between different physical activity (PA) intensities and cardiometabolic risk factors among a sample of Canadian youth. METHODS: The findings are based on a 2-year prospective cohort study in a convenience sample of 315 youth aged 9-15 years at baseline from rural and urban schools in Alberta, Canada. Different intensities (light, moderate and vigorous) of PA were objectively assessed with Actical accelerometers. The main outcome measures were body mass index (BMI) z-score, waist circumference, cardiorespiratory fitness and systolic blood pressure at 2-year-follow-up and conditional BMI z-score velocity. A series of linear regression models were conducted to investigate the associations after adjusting for potential confounders. RESULTS: At follow-up, cardiorespiratory fitness increased (quartile 1 vs quartile 4=43.3 vs 50.2; P(trend)<0.01) and waist circumference decreased (quartile 1 vs quartile 4=79.0 vs 72.6; P(trend)=0.04; boys only) in a dose-response manner across quartiles of baseline vigorous-intensity PA. A similar trend was observed for systolic blood pressure (quartile 1 vs quartile 4=121.8 vs 115.3; P(trend)=0.07; boys only). Compared with quartile 1 of vigorous-intensity PA, BMI z-score at follow-up and conditional BMI z-score velocity were significantly lower in the quartile 2 and 3 (P<0.05). Waist circumference at follow-up also decreased (quartile 1 vs quartile 4=75.3 vs 73.8; P(trend)=0.04) across quartiles of baseline moderate-intensity PA. CONCLUSIONS: Time spent in vigorous-intensity PA was associated with several positive health outcomes 2 years later. These findings suggest that high-intensity activities in youth help to reduce the risk for several chronic diseases.

Use of hyaluronan in the selection of sperm for intracytoplasmic sperm injection (ICSI): significant improvement in clinical outcomes--multicenter, double-blinded and randomized controlled trial.

STUDY QUESTION: Does the selection of sperm for ICSI based on their ability to bind to hyaluronan improve the clinical pregnancy rates (CPR) (primary end-point), implantation (IR) and pregnancy loss rates (PLR)? SUMMARY ANSWER: In couples where ≤ 65% of sperm bound hyaluronan, the selection of hyaluronan-bound (HB) sperm for ICSI led to a statistically significant reduction in PLR. WHAT IS KNOWN AND WHAT THIS PAPER ADDS: HB sperm demonstrate enhanced developmental parameters which have been associated with successful fertilization and embryogenesis. Sperm selected for ICSI using a liquid source of hyaluronan achieved an improvement in IR. A pilot study by the primary author demonstrated that the use of HB sperm in ICSI was associated with improved CPR. The current study represents the single largest prospective, multicenter, double-blinded and randomized controlled trial to evaluate the use of hyaluronan in the selection of sperm for ICSI. DESIGN: Using the hyaluronan binding assay, an HB score was determined for the fresh or initial (I-HB) and processed or final semen specimen (F-HB). Patients were classified as >65% or ≤ 65% I-HB and stratified accordingly. Patients with I-HB scores ≤ 65% were randomized into control and HB selection (HYAL) groups whereas patients with I-HB >65% were randomized to non-participatory (NP), control or HYAL groups, in a ratio of 2:1:1. The NP group was included in the >65% study arm to balance the higher prevalence of patients with I-HB scores >65%. In the control group, oocytes received sperm selected via the conventional assessment of motility and morphology. In the HYAL group, HB sperm meeting the same visual criteria were selected for injection. Patient participants and clinical care providers were blinded to group assignment. PARTICIPANTS AND SETTING: Eight hundred two couples treated with ICSI in 10 private and hospital-based IVF programs were enrolled in this study. Of the 484 patients stratified to the I-HB > 65% arm, 115 participants were randomized to the control group, 122 participants were randomized to the HYAL group and 247 participants were randomized to the NP group. Of the 318 patients stratified to the I-HB ≤ 65% arm, 164 participants were randomized to the control group and 154 participants were randomized to the HYAL group. MAIN RESULTS AND THE ROLE OF CHANCE: HYAL patients with an F-HB score ≤ 65% demonstrated an IR of 37.4% compared with 30.7% for control [n = 63, 58, P > 0.05, (95% CI of the difference -7.7 to 21.3)]. In addition, the CPR associated with patients randomized to the HYAL group was 50.8% when compared with 37.9% for those randomized to the control group (n = 63, 58, P > 0.05). The 12.9% difference was associated with a risk ratio (RR) of 1.340 (RR 95% CI 0.89-2.0). HYAL patients with I-HB and F-HB scores ≤ 65% revealed a statistically significant reduction in their PLR (I-HB: 3.3 versus 15.1%, n = 73, 60, P = 0.021, RR of 0.22 (RR 95% CI 0.05-0.96) (F-HB: 0.0%, 18.5%, n = 27, 32, P = 0.016, RR not applicable due to 0.0% value) over control patients. The study was originally planned to have 200 participants per arm providing 86.1% power to detect an increase in CPR from 35 to 50% at α = 0.05 but was stopped early for financial reasons. As a pilot study had demonstrated that sperm preparation protocols may increase the HB score, the design of the current study incorporated a priori collection and analysis of the data by both the I-HB and the F-HB scores. Analysis by both the I-HB and F-HB score acknowledged the potential impact of sperm preparation protocols. BIAS, CONFOUNDING AND OTHER REASONS FOR CAUTION: Selection bias was controlled by randomization. Geographic and seasonal bias was controlled by recruiting from 10 geographically unique sites and by sampling over a 2-year period. The potential for population effect was controlled by adjusting for higher prevalence rates of >65% I-HB that naturally occur by adding the NP arm and to concurrently recruit >65% and ≤ 65% I-HB subjects. Monitoring and site audits occurred regularly to ensure standardization of data collection, adherence to the study protocol and subject recruitment. Subgroup analysis based on the F-HB score was envisaged in the study design. GENERALIZABILITY TO OTHER POPULATIONS: The study included clinics using different sperm preparation methods, located in different regions of the USA and proceeded in every month of the year. Therefore, the results are widely applicable.

Feasibility, user experiences, and preliminary effect of Conversation Cards for Adolescents© on collaborative goal-setting and behavior change: protocol for a pilot randomized controlled trial.

BACKGROUND: Adolescents and providers can benefit from practical tools targeting lifestyle modification for obesity prevention and management. We created Conversation Cards for Adolescents© (CCAs), a patient-centered communication and behavior change tool for adolescents and providers to use in clinical practice. The purpose of our study is to (i) assess the feasibility of CCAs in a real-world, practice setting to inform full-scale trial procedures, (ii) assess user experiences of CCAs, and (iii) determine the preliminary effect of CCAs on changing behavioral and affective-cognitive outcomes among adolescents. METHODS: Starting in early 2019, this prospective study is a nested mixed-methods, theory-driven, and pragmatic pilot randomized controlled trial with a goal to enroll 50 adolescents (13-17 years old) and 9 physicians practicing at the Northeast Community Health Centre in Edmonton, Alberta, Canada. Adolescents will collaboratively set one S.M.A.R.T. (specific, measurable, attainable, realistic, timely) goal with their physician to implement over a 3-week period; however, only those randomized to the experimental group will use CCAs to inform their goal. Outcome assessments at baseline and follow-up (3 weeks post-baseline) will include behavioral, affective-cognitive, and process-related outcomes. DISCUSSION: In examining the feasibility, user experiences, and preliminary effect of CCAs, our study will add contributions to the obesity literature on lifestyle modifications among adolescents in a real-world, practice setting as well as inform the scalability of our approach for a full-scale effectiveness randomized controlled trial on behavior change. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03821896.

Defining metabolically healthy obesity in children: a scoping review.

We conducted a scoping review to identify definitions of metabolically healthy obesity (MHO), describe gaps in the literature, and establish a universal definition of MHO in children. We searched electronic databases from January 1980 to June 2017 and grey literature. Experimental, quasi-experimental, or observational studies were eligible for inclusion if they (i) included a definition of MHO that identified risk factors, cut-off values, and the number of criteria used to define MHO, and (ii) classified 2-18 year olds as overweight or obese. Two reviewers independently screened 1,711 papers for relevance and quality; we extracted data from 39 individual reports that met inclusion criteria. Most (31/39; 79%) definitions of MHO included an absence of cardiometabolic risk factors. Heterogeneity across MHO definitions, obesity criteria, and sample sizes/characteristics resulted in variable prevalence estimates (3-80%). Finally, we convened an international panel of 46 experts to complete a 4-round Delphi process to generate a consensus-based definition of MHO. Based on consensus (≥ 80% agreement), our definition of MHO included: high density lipoprotein-cholesterol > 40 mg/dl (or > 1.03 mmol/l), triglycerides ≤ 150 mg/dl (or ≤ 1.7 mmol/l), systolic and diastolic blood pressure ≤ 90th percentile, and a measure of glycemia. This definition of MHO holds potential universal value to enable comparisons between studies and inform clinical decision-making for children with obesity.

A brief eHealth tool delivered in primary care to help parents prevent childhood obesity: a randomized controlled trial.

OBJECTIVES: To determine the feasibility and preliminary impact of an electronic health (eHealth) screening, brief intervention and referral to treatment (SBIRT) delivered in primary care to help parents prevent childhood obesity. METHODS: Parents of children (5-17 years) were recruited from a primary care clinic. Children's measured height and weight were entered into the SBIRT on a study-designated tablet. The SBIRT screened for children's weight status, block randomized parents to one of four brief interventions or an eHealth control and provided parents with a menu of optional obesity prevention resources. Feasibility was determined by parents' interest in, and uptake of, the SBIRT. Preliminary impact was based on parents' concern about children's weight status and intention to change lifestyle behaviours post-SBIRT. RESULTS: Parents (n = 226) of children (9.9 ± 3.4 years) were primarily biological mothers (87.6%) and Caucasian (70.4%). The proportion of participants recruited (84.3%) along with parents who selected optional resources within the SBIRT (85.8%) supported feasibility. Secondary outcomes did not vary across groups, but non-Caucasian parents classified as inaccurate estimators of children's weight status reported higher levels of concern and intention to change post-SBIRT. CONCLUSIONS: Our innovative, eHealth SBIRT was feasible in primary care and has the potential to encourage parents of unhealthy weight children towards preventative action.

Barriers to and enablers of healthy lifestyle behaviours in adolescents with obesity: a scoping review and stakeholder consultation.

Healthy lifestyle behaviours are key to successful weight management, but have proven to be challenging to attain for adolescents with obesity. The purpose of our scoping review was to (i) describe barriers and enablers that adolescents with obesity encounter for healthy nutrition, physical activity, sedentary behaviour and sleep habits and (ii) identify gaps in the literature. We adhered to established methodology for scoping reviews. Six databases were searched (1980-June 2016) for original articles published in English or French that focused on lifestyle behaviours of 13- to 17-year-olds in paediatric weight management. Following screening and data extraction, findings of selected articles were synthesized thematically using a social ecological framework. Stakeholder consultation (n = 20) with adolescents with obesity and health professionals was completed to enhance methodological rigour. Our search yielded 17 articles for inclusion, including 546 unique participants. Barriers to healthy nutrition and physical activity were more consistently related to individual-level and interpersonal-level factors; enablers tended to be linked with interpersonal-level factors. Knowledge gaps identified related to sedentary behaviour and sleep as well as environmental and policy levels of influence. Our review revealed that some barriers and enablers were unique to adolescents with obesity, which were either within or beyond their control. These findings highlight the importance of multilevel interventions to enable healthy lifestyle behaviours for weight management.

Communicating with children and families about obesity and weight-related topics: a scoping review of best practices.

BACKGROUND: Healthcare professionals have called for direction on how best to communicate about weight-related topics with children and families. Established scoping review methodology was used to answer the question: 'How can healthcare professionals best communicate with children and their families about obesity and weight-related topics?' METHODS: We searched four scientific databases, two grey literature repositories and 14 key journals (2005-2016). Inclusion criteria were (i) children up to and including 18 years of age and/or their parents; (ii) communication about healthy weight, overweight, obesity or healthy/active living; and (iii) healthcare setting. RESULTS: Thirty-two articles were included. Evidence-based best practices were largely absent from the literature, although the following guiding principles were identified: (i) include all stakeholders in discussions; (ii) raise the topic of weight and health early and regularly; (iii) use strengths-based language emphasizing health over weight; (iv) use collaborative goal-setting to engage children and parents and (v) augment discussions with appropriate tools and resources. Guidance on how to implement these principles and how to negotiate relevant contextual factors (e.g. age, culture and disability) is still needed. CONCLUSION: Despite agreement on a number of guiding principles, evidence-based weight-related communication best practices are lacking. Rigorous, empirical evaluations of communication approaches are urgently required, especially those that include children's perspectives.

Variations in the prevalence and predictors of prevalent metabolically healthy obesity in adolescents.

BACKGROUND: Obesity is a heterogeneous condition, which includes a subset of individuals that can be classified as having metabolically healthy obesity (MHO), but there is no consensus on what constitutes MHO. Thus, the objective of the study is to examine the prevalence and predictors of prevalent MHO in adolescents using various definitions of MHO. METHODS: Cross-sectional data from the 1999-2010 National Health and Nutrition Examination Surveys were used. Participants included 316 male and 316 female adolescents aged 12-19 years with a BMI ≥ 95th percentile. Two definitions were used to define MHO. First, MHO was defined as having ≤1 metabolic syndrome criteria (excluding waist) and being free of type 2 diabetes, hypertension and dyslipidemia. Second, MHO was defined as being free of all metabolic syndrome criteria, insulin resistance and inflammation. RESULTS: The prevalence of MHO was 42% (male) and 74% (female) using the first definition and 7% (male) and 12% (female) using the second more conservative definition. Lower abdominal obesity (waist circumference) and lower insulin resistance predicted prevalent MHO in male and female adolescents for both definitions (p < 0.01). Associations between dietary components and MHO were weak and inconsistent, while physical activity and inflammation were not associated with MHO in male and female adolescents for both definitions (p > 0.05). CONCLUSIONS: The prevalence of MHO in adolescents varied across definitions, with lower levels of abdominal obesity and insulin resistance as the most consistent predictors of prevalent MHO status.

Weight bias reduction in health professionals: a systematic review.

Innovative and coordinated strategies to address weight bias among health professionals are urgently needed. We conducted a systematic literature review of empirical peer-reviewed published studies to assess the impact of interventions designed to reduce weight bias in students or professionals in a health-related field. Combination sets of keywords based on three themes (1: weight bias/stigma; 2: obesity/overweight; 3: health professional) were searched within nine databases. Our search yielded 1447 individual records, of which 17 intervention studies satisfied the inclusion criteria. Most studies (n = 15) included medical, dietetic, health promotion, psychology and kinesiology students, while the minority included practicing health professionals (n = 2). Studies utilized various bias-reduction strategies. Many studies had methodological weaknesses, including short assessment periods, lack of randomization, lack of control group and small sample sizes. Although many studies reported changes in health professionals' beliefs and knowledge about obesity aetiology, evidence of effectiveness is poor, and long-term effects of intervention strategies on weight bias reduction remain unknown. The findings highlight the lack of experimental research to reduce weight bias among health professionals. Although changes in practice will likely require multiple strategies in various sectors, well-designed trials are needed to test the impact of interventions to decrease weight bias in healthcare settings.

Ready, set, go! Motivation and lifestyle habits in parents of children referred for obesity management.

BACKGROUND: Parents play a fundamental role in helping children with obesity to make and maintain healthy lifestyle changes. OBJECTIVE: This study aimed to characterize stages of engagement to change nutrition and physical activity habits among parents whose children with obesity were enrolled in obesity management and examine differences in parents' own nutrition and physical activity habits according to their stage of engagement. METHODS: Medical records of 113 children (body mass index [BMI] ≥95th percentile) enrolled in an outpatient weight management clinic were reviewed for baseline (cross-sectional) data. Parents completed the Weight Loss Behavior-Stage of Change Scale to assess the degree of engagement in making healthy changes to their lifestyle behaviours. Latent class analysis was used to classify parents into distinct clusters by grouping individuals with similar ratings of stages of engagement regarding nutrition- and physical activity-related behaviours. RESULTS: Parents' engagement in healthy lifestyle behaviours varied (more engaged [n = 43]; less engaged [n = 70]). A greater proportion of parents in the more engaged group was in action and/or maintenance stages of changing their lifestyle habits. The more engaged group was less overweight than the less engaged group (BMI = 28.5 vs. 33.3 kg m(-2); P < 0.05). Further, the more engaged group consumed fewer total calories, calories from fat, trans fat and carbohydrates vs. their less engaged peers (P < 0.05). Compared with the less engaged group, the more engaged group consumed more daily servings of vegetables and fruits (4.9 vs. 3.9, P < 0.05) and accumulated more steps per day (9130 vs. 7225; P < 0.05). The more engaged group was also more likely to meet daily recommendations for vegetable and fruit intake (48.8 vs. 24.3%; P < 0.05) and physical activity (42.9 vs. 22.9%, P < 0.05). CONCLUSIONS: Parents of children with obesity varied in their degree of engagement in making healthy changes to their lifestyle behaviours, and those categorized as more engaged already demonstrated positive lifestyle behaviours. Information regarding parents' degree of engagement in healthy behaviours can inform clinical recommendations, especially when parents represent the primary agents of change in families trying to manage paediatric obesity.

Glycosaminoglycans in bovine cumulus-oocyte complexes: morphology and chemistry.

Bovine cumulus-oocyte complexes from small (1-5 mm) follicles were cultured for 24 h in 0.25 ml minimum essential medium supplemented with 10% fetal bovine serum and 20 microCi [3H]glucosamine. Treatment groups consisted of supplementing the culture medium with no hormone (control), 0.5 IU/ml follicle-stimulating hormone (FSH) or 10 mM 8-Br-adenosine cyclic monophosphate (cAMP). After culture, the complexes were fixed for light and scanning electron microscopy. Electron photomicrographs revealed that complexes induced to expand with FSH or cAMP contained a copious glycosaminoglycan (GAG) matrix extending between and around the cumulus cells. Control complexes did not exhibit expansion or an extracellular matrix. The radiolabeled GAG material was isolated for chemical identification. Chemical analyses included: (1) electrophoresis of GAG material, (2) electrophoresis of GAG material after enzyme or nitrous acid treatment, (3) thin-layer chromatography of GAG hydrolysates. The results from electrophoresis showed that the radiolabeled GAG co- migrated with hyaluronic acid. The GAG material was resistant to chondroitinase ABC and nitrous acid degradation but was digested by hyaluronidase. Complexes treated with FSH and cAMP incorporated higher (P less than 0.1 and P less than 0.025 respectively) amounts of [3H]glucosamine into hyaluronic acid than control cultures. Thin-layer chromatography identified the primary amino sugar of the GAG to be glucosamine. These data collectively showed that the radioactive GAG produced by bovine cumulus-oocyte complexes was hyaluronic acid.

Characterization of human sperm antigens and antisperm antibodies in infertile patients.

OBJECTIVE: To identify which sperm antigens may elicit the production of functionally important antisperm antibodies. DESIGN: Immunoblot analysis was performed on 69 serum and 9 seminal plasma samples from infertile patients, using detergent extracts of pooled donor sperm as the antigen source. Serum and seminal plasma had been previously tested by an indirect immunobead binding test (IBT); 61 IBT-positive and 17 IBT-negative samples were included in the study. Proteins recognized by IBT-positive but not IBT-negative samples were most likely to be cell surface antigens, whereas proteins recognized by both IBT-positive and IBT-negative samples were probably intracellular. Antibodies directed toward surface antigens would be most likely to affect fertilization. Characterization of sperm surface proteins on both acrosome-intact and -reacted sperm used labeling of cell surface proteins with an N-hydroxysuccinimide ester of biotin, fractionation of sperm heads and tails, and lectin binding to determine glycosylation. RESULTS: Specific immunoreactivity (with respect to IBT results) was observed to 35K, 40 to 45K, 57K, 66K, and 88 to 90K MW proteins. Characterization studies identified an 88K MW glycosylated plasma membrane protein, a 66K MW inner acrosomal membrane protein, a 34K MW inner acrosomal membrane protein, and a 35K MW prominent tail protein. CONCLUSION: Immunological infertility may involve several antigens characterized in this study. Further studies are necessary to determine if antibodies to these specific proteins interfere with sperm function.

Cryopreservation of human spermatozoa. II. Postthaw chronology of motility and of zona-free hamster ova penetration.

Postthaw dynamics of motility maintenance and ability to penetrate zona-free hamster ova were examined with human sperm. Ten semen samples were each divided into two equal volumes; one was cryopreserved while the other half remained untreated. Frozen samples were thawed, and initial evaluations for motility and hamster egg penetration were made on both untreated and frozen-thawed samples. The time difference between the initial evaluations for the two treatment groups was approximately 30 minutes as a result of the time required to freeze and thaw aliquots. Subsequent evaluations were made 6, 12, 24, and 48 hours later. Over all times both the motility and fertilizability of cryopreserved spermatozoa were significantly reduced (P less than 0.05) when compared with those of untreated sperm. The pattern of motility loss over time was similar between untreated and frozen-thawed sperm (P greater than 0.10). Conversely, differences between untreated and frozen-thawed sperm in fertilizability patterns were dramatic (P less than 0.05). This was evidenced by penetration rates for cryopreserved sperm highest at 0 hour and decreasing over time, whereas penetration by untreated spermatozoa was lowest at 0 hour, increasing to a maximum at 24 hours. These observations may be important in the development of laboratory protocols for freezing and clinical protocols for using frozen-thawed sperm.

Cryopreservation of human spermatozoa. III. The effect of cryoprotectants on motility.

A series of experiments was conducted to examine potential toxic effects of cryoprotectants on motility of human spermatozoa. The data indicated that exposure of spermatozoa to cryoprotectant medium for as little as 15 minutes at room temperature caused a reduction in motility. This reduction in motility was caused by glycerol. Lowering glycerol concentrations from 7.5% to 5.0% improved sperm motility at 24 hours post-thaw. Sperm motility was not affected by either slow or abrupt cooling rates above -5 degrees C. Motility was greater in cryopreserved sperm at 24 hours post-thaw when glycerol was added at -5 degrees C rather than at room temperature. These data suggest that avoiding glycerol toxicity either by reducing the concentration used or by adding glycerol at a lower temperature, or both, may improve human sperm cryosurvival rates.

Cryopreservation of hamster oocytes: effects of vitrification or freezing on human sperm penetration of zona-free hamster oocytes.

Three experiments were conducted for evaluation of the efficacy of conventional freezing or vitrification of hamster oocytes for use in a human sperm penetration assay (hSPA). In experiment 1, oocytes were cryopreserved and evaluated for survival on the basis of morphologic criteria. Survival of vitrified oocytes and that of frozen oocytes were not different, whereas all cryopreserved groups had lower survival than noncryopreserved controls. In experiment 2, oocytes were conventionally frozen or vitrified and evaluated in an hSPA. Vitrified oocytes had a lower frequency of sperm penetration than frozen oocytes, and all cryopreserved groups had lower penetration rates than untreated controls. In experiment 3, oocytes were exposed to the cryoprotectant used to vitrify (VS1) or freeze (DMSO) but not cooled prior to evaluation in an hSPA. Exposure to DMSO but not VS1 reduced hSPA values. It is concluded from these experiments that while all cryopreserved oocytes do not survive, at current stages of development conventionally frozen oocytes perform better than vitrified oocytes in the hSPA and losses associated with conventional freezing procedures may be related to cryoprotectant exposure, whereas vitrification losses are more probably due to events associated with rapid cooling and/or warming of the oocytes.

Cryopreservation of human spermatozoa. I. Effects of holding procedure and seeding on motility, fertilizability, and acrosome reaction.

Three experiments were conducted to evaluate effects of holding semen at +5.0 degrees C for 30 minutes or -5.0 degrees C for 10 minutes and ice crystal induction (seeding) on frozen-thawed human spermatozoa. In experiment 1, spermatozoa were frozen, and postthaw motility was evaluated immediately (0 hour) and 24 hours later. At both 0 and 24 hours, nonfrozen control samples had higher motility than all other treatment groups. At 0 hour postthaw, motility was higher in samples held at -5.0 degrees C for 10 minutes with no significant effect of seeding. At 24 hours, samples held at -5.0 degrees C for 10 minutes and seeded, but not samples held at -5.0 degrees C and not seeded, had higher motility than samples held at +5.0 degrees C. In experiment 2, semen samples were frozen, and fertilizability was evaluated in a zona-free hamster egg penetration assay. Seeded samples had a higher frequency of sperm penetration than either nonfrozen or nonseeded samples. In experiment 3, nonfrozen controls and frozen treatment groups were evaluated for the frequency of survival and acrosomal integrity. Seeded samples had higher frequencies of survival and loss of acrosomal integrity than nonseeded samples. All frozen-thawed samples had a lower frequency of survival and a higher frequency of loss of acrosomal integrity than nonfrozen controls. Although altered patterns of fertilizability and acrosomal integrity are induced, collectively these data suggest that incorporating a holding temperature of -5.0 degrees C for 10 minutes and seeding may result in a superior protocol for freezing human spermatozoa.