Acellularization of human placenta with preservation of the basement membrane: a potential matrix for tissue engineering

Basement membrane directs cell migration, cell adherence, and tissue organization in vivo. These qualities would be advantageous in tissue engineering, during which repopulation of scaffolds by functional cells is a common problem. The purpose of this study was to expose human term placenta to a chemical acellularization protocol using the isolated cotyledon perfusion technique to develop an acellular scaffold rich in extracellular matrix components for tissue engineering applications. The processed blocks of tissue demonstrated no cellular material on histologic examination. Periodic acid-Schiff staining, type IV collagen labeling, and transmission electron microscopy demonstrated preservation of the extracellular matrix and basement membrane. Maintenance of the extracellular matrix architecture was seen on scanning electron microscopy. Placenta-derived acellular matrix is a potential volume scaffold for tissue engineering. It is rich in extracellular matrix, has a complex architecture of vascular channels, and has unique donor opportunities.     

A treadmill ramp protocol using simultaneous changes in speed and grade

INTRODUCTION: A treadmill exercise test requiring a low initial metabolic rate that then increments the work rate linearly to reach the subject's limit of tolerance in approximately 10 min would have significant advantages for exercise testing and rehabilitation of subjects with impaired exercise tolerance. METHODS: We developed such a treadmill protocol that uses a linear increase in walking speed coupled with a curvilinear increase in treadmill grade to yield a linear increase in work rate. RESULTS: Twenty-two healthy, sedentary subjects performed both this new treadmill protocol and a standard cycle ergometry ramp protocol eliciting similar work rate profiles. The low initial treadmill speed and grade resulted in a low initial metabolic rate, commensurate with unloaded pedaling on a cycle ergometer (average [OV0312]O2 = 0.54 +/- 0.16 vs 46 +/- 0.12 l x min(-1)). This combination of simultaneous increase in speed and grade yielded a linear work rate and its oxygen uptake response (R2 = 0.96 +/- 0.03) with a slope of 11.4 +/- 2.4 ml x min(-1) x W(-1)-slightly, but significantly, higher than on the cycle (9.6 +/- 2.0 ml x min(-1) x W(-1)). This difference was attributed to unmeasured work associated, for example, with additional limb movements and frictional losses. As previously demonstrated, both the peak oxygen uptake and the estimated lactate threshold were higher on the treadmill than for cycle ergometry (averaging 23% and 27%, respectively, in these subjects). CONCLUSION: This treadmill protocol provides a linear profile of work rate as is currently standard for cycle ergometry and is appropriate for testing of subjects with low exercise tolerance.     

Women awaiting knee replacement have reduced function and growth hormone

Women experience greater functional impairment and occurrence of osteoarthritis than men. We hypothesized that lower levels of serum insulin-like growth factor-I may contribute to gender differences in physical impairments. A cross-sectional comparison (n = 139) was done on candidates having total knee arthroplasty and healthy controls subjects of similar age (range, 55-75 years). Physical function, perceived function, and serum insulin-like growth factor levels were compared across group and gender using analysis of variance. Insulin-like growth factor-I values were markedly reduced in women overall and women having surgery had significantly reduced levels despite lean body mass correction. Values consistent with clinical hormone deficiency were observed in 21% of women and only 4% of men having arthroplasty. Physical function was markedly reduced in women, at times functioning only 33% when compared with healthy women, whereas men's limitations were not as profound. The current findings indicate that physical function is more impaired and serum insulin-like growth factor-I is markedly reduced in women awaiting arthroplasty than their male counterparts. The gender differences observed biochemically and with functional performance indicate that the pathophysiology of end stage osteoarthritis may differ between men and women.     

Reparative changes and the false-positive/false-negative Papanicolaou test: a study from the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology

CONTEXT: Reparative changes can be a diagnostic challenge on Papanicolaou tests and must be distinguished from epithelial cell abnormalities. Both squamous intraepithelial lesions (SIL) and carcinoma may be underdiagnosed as repair. This study examines laboratory and cytologist performance in the diagnosis of repair. OBJECTIVES: To determine if laboratories and cytologists can consistently distinguish reparative changes from SIL and carcinoma and to document how often SIL and carcinoma are mistaken for repair in a standardized educational slide program. DESIGN: Results for reparative changes, SIL, and carcinoma slides from the College of American Pathologists Interlaboratory Comparison Program in Cervicovaginal Cytology for 1998 were analyzed. Only results from validated referenced slide responses were used. RESULTS: The concordancy rate for reparative-change slides, that is, any response of within normal limits or benign cellular changes, ranged from 91% to 94% for cytotechnologist, pathologist, and laboratory responses. False-negative rates for squamous carcinoma and adenocarcinoma, high-grade SIL, and low-grade SIL ranged from 0.47% to 5.41%; the proportion of false-negative diagnoses of reparative changes ranged from 24% to 62% of all discordant responses. CONCLUSIONS: Of all benign cellular changes and within normal limits categories in the Interlaboratory Comparison Program in Cervicovaginal Cytology, repair most often elicits a false-positive laboratory response. Underdiagnosing epithelial abnormalities as repair is also a source of false-negative Papanicolaou test results.     

Skeletal growth and long-term bone turnover after enterocystoplasty in a chronic rat model

OBJECTIVE: To investigate skeletal growth and bone metabolism in a chronic animal model of urinary diversion. MATERIALS AND METHODS: Young male Wistar rats (120) were allocated randomly to four groups undergoing: ileocystoplasty, ileocystoplasty and resection of the ileocaecal segment, colocystoplasty, and controls. All animals received antibiotics for 1 week after surgery; half of each group remained on oral antibiotics. Bone-related biochemistry was measured in serum and urine. Dual-energy X-ray absorptiometry and peripheral quantitative computed tomography (pQCT) were used to determine bone mass ex vivo. RESULTS: Most (90%) of the rats survived the study period (8 months); six rats died from bowel obstruction at the level of the entero-anastomosis and four had to be killed because of persistent severe diarrhoea. Vital intestinal mucosa was found in all augmented bladders. There were no differences in bone length and volume. Loss of bone mass was almost exclusively in rats with ileocystoplasty and resection of the ileocaecal segment (-37.5%, pQCT, P < 0.01). There was no hyperchloraemic metabolic acidosis or gross impairment of renal function. Hypomagnesaemia, hypocalcaemia and decreased insulin-like growth factor-binding protein 3 were the only significant findings on blood analysis. Deoxypyridinoline crosslinks in urine were higher in rats with an enterocystoplasty than in controls. CONCLUSIONS: Enterocystoplasty in rats neither impairs skeletal growth nor bone quantity, but leads to significant loss of bone mass when combined with resection of the ileocaecal segment. Rarefaction of the trabecular network is confined to the metabolically highly active cancellous compartment, most likely as a consequence of intestinal malabsorption.     

Enterocystoplasty in childhood: a second look at the effect on growth

OBJECTIVE: To re-evaluate the assumption that enterocystoplasty in children has a detrimental effect on linear growth (which is almost exclusively based upon a chance finding in a retrospective study 10 years ago) in a larger cohort and with a longer follow-up. PATIENTS AND METHODS: The original 12 children who had impaired linear growth in a previous study 10 years earlier were re-measured. A larger cohort was identified from the 242 children and adolescents who had undergone enterocystoplasty between 1982 and 1997. Patients with conditions involving organ systems apart from the urinary tract, and those with myelomeningocele, malignant diseases, reduced glomerular filtration rate and incomplete notes were excluded. In the definitive study cohort (123; mean age at operation 8.6 years; mean age at investigation 16.8 years) enterocystoplasty had been undertaken using colon in 70, ileum in 37, a combination of both in 11, ileocaecal segments in three and stomach in two patients. RESULTS: Of the original 12 patients, six had regained or surpassed their preoperative position on their growth charts. In all patients with a known target centile range the final height was within their genetic growth potential. In the cohort of 123 patients, 1215 height and weight measurements had been recorded. The distribution of percentile positions before and after enterocystoplasty showed a normal configuration, with 83% and 80% of patients growing within two standard deviations of the 50th percentile. After surgery, 85% either remained on the same or reached a higher centile. Nineteen (15.5%) were in a lower position, with a similar tendency in the weight centile. A clinically relevant growth disorder was recognized in four patients with a complete endocrinological evaluation; in none of these was enterocystoplasty thought to be a causal factor. CONCLUSIONS: It is very unlikely that the loss of the preoperative percentile position on the growth curve in 15% of children after enterocystoplasty is a consequence of the surgery. Rather it is a non-specific phenomenon that has to be considered in any clinical population of the same size and age distribution after the same length of time.     

The effect of enterocystoplasty on bone strength assessed at four different skeletal sites in a rat model

The objective of the study was to investigate bone strength at four different skeletal sites in a chronic animal model of urinary diversion. Young male Wistar rats (120) were allocated randomly to four groups undergoing ileocystoplasty; ileocystoplasty and resection of the ileocecal segment; colocystoplasty; or sham operation (controls). After 8 months the lumbar vertebrae, femora, and tibiae were harvested at necropsy. Bone strength was assessed biomechanically at four different skeletal sites: vertebra L3, femoral middiaphysis, femoral neck, and distal femoral metaphysis. Bone mass and architecture were assessed using standard static histomorphometry of the proximal tibial metaphysis (trabecular bone volume [BV/TV]; trabecular number [Tb.N]) and ash weight. Statistically significant differences of biomechanical parameters between groups were observed at three skeletal sites with corresponding changes in tibial histomorphometry. Isolated ileocystoplasty resulted in decreased maximum load values of L3 (-16.4%; p < 0.0035) and a substantial reduction in tibial BV/TV (-34.7%; p < 0.05). Ileocystoplasty combined with resection of the ileocecal segment led to a significant loss of bone strength of L3 (-32.4%; p < 0.0015) and a dramatic reduction of tibial BV/TV (-45.9%; p < 0.01). Loss of tibial metaphyseal bone mass was predominantly caused by a decrease in Tb.N. (p < 0.01). Colonic augmentation had no significant effect on bone strength or histomorphometric values. In conclusion, this is the first experimental study to demonstrate the relevance of histomorphometrically proven bone loss after enterocystoplasty in terms of biomechanical variables.     

Exchangeable zinc pool masses and turnover are maintained in healthy men with low zinc intakes

Previous studies suggest that rapidly exchanging zinc pools (EZP), thought to supply the zinc required by tissues, are smaller and turn over more rapidly in individuals with lower zinc intakes. We studied the effects of low dietary zinc (4.6 mg/d) on EZP mass and turnover in seven healthy men confined during a 20-wk clinical study. Supplements of 9.1 mg zinc were given during the 5-wk baseline and repletion periods, and placebos were given during a 10-wk zinc-restriction period. Stable 70Zn tracers were administered intravenously at the end of baseline, 3 and 10 wk after the start of zinc restriction and at the end of repletion. Multiple plasma samples were collected over an 8-d period after tracer administration. 70Zn:66Zn ratios were measured using inductively coupled plasma mass spectrometry, and tracer-tracee data were analyzed by compartmental modeling. Activities of the zinc-dependent enzymes, alkaline phosphatase and 5'nucleotidase, were unchanged during the study. There were no significant changes in EZP masses or kinetic parameters. A three-compartment model indicated that the masses of plasma zinc and total EZP averaged 3.25 +/- 0.58 and 147.8 +/- 33.2 mg, respectively, at the four time points studied. Plasma zinc mass turned over at an average of 5.3 times per hour. There was an 11% reduction (P = 0.06) in plasma zinc flux 3 wk after the start of the low zinc diet period, but it returned to baseline values after 10 wk of zinc restriction. The results suggest that total EZP mass is maintained when dietary zinc is reduced to 4.6 mg/d over a 10-wk period.