On the regeneration of fish scales: structure and mechanical behavior.

Fish scales serve as a dermal armor that provides protection from physical injury. Owing to a number of outstanding properties, fish scales are inspiring new concepts for layered engineered materials and next-generation flexible armors. Although past efforts have primarily focused on the structure and mechanical behavior of ontogenetic scales, the structure-property relationships of regenerated scales have received limited attention. In the present study, common carp (Cyprinus carpio) acquired from the wild were held live in an aquatic laboratory at 10°C and 20°C. Ontogenetic scales were extracted from the fish for analysis, as well as regenerated scales after approximately 1 year of development and growth. Their microstructure was characterized using microscopy and Raman spectroscopy, and the mechanical properties were evaluated in uniaxial tension to failure under hydrated conditions. The strength, strain to fracture and toughness of the regenerated scales were significantly lower than those of ontogenetic scales from the same fish, regardless of the water temperature. Scales that regenerated at 20°C exhibited significantly higher strength, strain to fracture and toughness than those regenerated at 10°C. The regenerated scales exhibited a highly mineralized outer layer, but no distinct limiting layer or external elasmodine; they also possessed a significantly lower number of plies in the basal layer than the ontogenetic scales. The results suggest that a mineralized layer develops preferentially during scale regeneration with the topology needed for protection, prior to the development of other qualities.

Clinical predictors of inflammatory bowel disease in a genetically well-defined Caucasian population.

BACKGROUND: Crohn's disease (CD) and ulcerative colitis (UC), the two main types of inflammatory bowel disease (IBD), are multifactorial conditions of unknown etiology. The objective of this study is to examine the combined gene-environment interactions influencing IBD susceptibility in a well-defined Caucasian cohort in rural mid-America. METHODS: Patients were diagnosed to have CD or UC using conventional radiologic, endoscopic, and/or histopathologic findings. Histological diagnosis was made by a single specialist gastrointestinal pathologist with a particular interest in IBD. Information regarding cigarette smoke exposure was obtained by administration of the Behavioral Risk Factor Surveillance System Survey (BRFSS) to all patients. Genomic DNA was extracted from peripheral blood leukocytes, and polymerase chain reaction (PCR) amplification and genotyping were performed for 11 Single Nucleotide Polymorphisms (SNP) in NOD2, IL23r, OCTN1 genes along with IGR. RESULTS: Our cohort consists of 1196 patients: 435 controls, 485 CD patients, and 276 UC patients. Only patients with genotype data for at least 7 of 11 SNPs were included in our data analysis. The control groups for all 11 SNPs were in Hardy-Weinberg Equilibrium. In genotype-association SNP analysis, all NOD2 SNPs (rs5743293, rs2066844, rs2066845) and the IL23r SNP (rs11465804) showed a significant association to IBD (p < 0.03). A multiple gene-interaction analysis showed an association between NOD2 and IL23r with UC (p = 0.04). There were no associations between any OCTN1 and IGR SNPs and IBD in this cohort. A multivariable logistic regression analysis showed that female gender, "current" or "former" smoking status, family history of IBD, and NOD2 SNP minor alleles were associated with CD. CONCLUSION: IBD remains to be challenging to properly diagnose, characterize, and treat. Our study proposes a combined genetic, phenotypic, and environmental approach in an attempt to better understand IBD. Previously demonstrated associations between OCTN1 and IGR and IBD were not confirmed.

Crohn's disease in Caucasians and African Americans, as defined by clinical predictors and single nucleotide polymorphisms.

OBJECTIVES: To compare three aspects of Crohn's disease (CD) between African Americans and Caucasians: (1) demographic data and environmental factors affecting CD susceptibility, (2) disease presentation and clinical course, and (3) genetic susceptibility via the use of single nucleotide polymorphism (SNP) data for inflammatory bowel disease (IBD) susceptibility loci. METHODS: Clinical data and peripheral blood were obtained from 1032 patients (554 CD patients and 478 controls) derived from a clinically well-defined university-based medical and surgical digestive disease practice and included those who were diagnosed with IBD. Genomic DNA was extracted and polymerase chain reaction (PCR) amplification and genotyping were performed for 11 SNPs, including the NOD2, IL-23r, OCTN 1, and the IGR gene variants. RESULTS: A total of 554 patients with CD were included in this study: 53 African Americans (10%), 485 Caucasians (87%), and 15 of other races (3%). The strongest demographic predictor of CD in African American patients was a family history of IBD. Ileocolic disease (L3) was the most common site involved in both African Americans and Caucasians, while the penetrating phenotype (B3) was the most common CD disease behavior in both races. Genotype association analysis showed a significant association between 2 IL23r gene SNPs and CD susceptibility in African Americans (p = .016 and .028, respectively). CONCLUSION: We believe this study is the first to report on genotype-phenotype associations in African American CD patients and compare findings to Caucasian CD patients within the same geographic area. We found no association between NOD2 gene SNPs and CD susceptibility in African Americans patients (p > .05).

Contributions of intermolecular bonding and lubrication to the mechanical behavior of a natural armor.

Among many dermal armors, fish scales have become a source of inspiration in the pursuit of "next-generation" structural materials. Although fish scales function in a hydrated environment, the role of water and intermolecular hydrogen bonding to their unique structural behavior has not been elucidated. Water molecules reside within and adjacent to the interpeptide locations of the collagen fibrils of the elasmodine and provide lubrication to the protein molecules during deformation. We evaluated the contributions of this lubrication and the intermolecular bonding to the mechanical behavior of elasmodine scales from the Black Carp (Mylopharyngodon piceus). Scales were exposed to polar solvents, followed by axial loading to failure and the deformation mechanisms were characterized via optical mechanics. Displacement of intermolecular water molecules by liquid polar solvents caused significant (p ≤ 0.05) increases in stiffness, strength and toughness of the scales. Removal of this lubrication decreased the capacity for non-linear deformation and toughness, which results from the increased resistance to fibril rotations and sliding caused by molecular friction. The intermolecular lubrication is a key component of the "protecto-flexibility" of scales and these natural armors as a system; it can serve as an important component of biomimetic-driven designs for flexible armor systems. STATEMENT OF SIGNIFICANCE: The natural armor of fish has become a topic of substantial scientific interest. Hydration is important to these materials as water molecules reside within the interpeptide locations of the collagen fibrils of the elasmodine and provide lubrication to the protein molecules during deformation. We explored the opportunity for tuning the mechanical behavior of scales as a model for next-generation engineering materials by adjusting the extent of hydrogen bonding with polar solvents and the corresponding interpeptide molecular lubrication. Removal of this lubrication decreased the capacity for non-linear deformation and toughness due to an increase in resistance to fibril rotations and sliding as imparted by molecular friction. We show that intermolecular lubrication is a key component of the "protecto-flexibility" of natural armors and it is an essential element of biomimetic approaches to develop flexible armor systems.

Characterization of embryonic antigens in the plasma of developing chick embryos.

Two prominent alpha-fetoproteins are found in chick embryo plasma up to and slightly beyond hatching. Only antisera against chick embryo plasma resolve them. alpha3-fetoprotein is a lipoprotein, the first fetoprotein thus described. It probably functions to transport lipid from the yolk to the embryo. The other fetoprotein (alpha4) is a major glycoprotein. Both are made by the liver and yolk sac. From 9 days onward, albumin is the principal export protein of the liver; at 7 days, when it first appears in the plasma, the yolk sac, but not the liver, makes it. The reverse situation occurs at 9 days. These facts suggest a humoral relationship between the yolk sac and liver. The embryo synthesizes every plasma protein including fetoproteins and "adult" proteins (prealbumin, albumin, and transferrin). No "cold" or unlabeled proteins are seen.

[Chemical and effect-related analysis of dioxins and dioxinlike PCB's]. / Chemische und wirkungsbezogene Analytik der Dioxine und dioxinähnlichen PCB.

Dioxins and dioxinlike PCBs are high toxic and persistent compounds. They accumulate in the feed and food chain. Because of their occurrence in a very low concentration range, the established requirements for analytical methods are restrictive. Commission Directive 2002/69/EC of 26 July 2002 lays down sampling methods and methods of analysis for the official control of dioxins and dioxinlike PCBs in foodstuff. Confirmatory methods are high resolution gas chromatography/high resolution mass spectrometry (HRGC/HRMS). Screening methods could be used to select samples with significant levels of dioxins. Bioassay--for example the EROD-Bioassay--can be used as a screening method.

Comparative outcomes of T-cell-depleted and non-T-cell-depleted allogeneic bone marrow transplantation for chronic myelogenous leukemia: impact of donor lymphocyte infusion.

PURPOSE: Donor lymphocyte infusion (DLI) can restore complete remission in patients with chronic myelogenous leukemia (CML) who have relapsed after T-cell-depleted (TCD) allogeneic bone marrow transplantation (BMT). The existence of salvage treatment for patients with DLI after TCD allogeneic BMT prompted an evaluation of overall outcome after CD6+ -TCD allogeneic BMT for patients treated during the time when DLI has been available. PATIENTS AND METHODS: We performed a retrospective analysis of outcomes of 46 patients who underwent TCD allogeneic BMT for stable-phase CML and compared these outcomes with those of 40 patients who underwent non-TCD allogeneic BMT. All subjects were patients at one of two neighboring institutions during a period when DLI was available. All patients received marrow from HLA-identical sibling donors, underwent similar myeloablative regimens, and had similar pretreatment characteristics. RESULTS: After BMT, the TCD group had a lower incidence of grade 2 to 4 acute (15% v 37%, P = .026) and chronic graft-versus-host disease (GVHD) (18% v 42%, P = .024) than did the non-TCD group. The 1-year treatment-related mortality rates for the TCD group and the non-TCD group were 13% and 29%, respectively (P = .07). The estimated 3-year probability of relapse (cytogenetic or hematologic) was higher for patients in the TCD group than for patients in the non-TCD group (62% v 24%, P = .0003). Twenty-three patients (20 in the TCD group and three in the non-TCD group) received and were assessable for response to DLI. After DLI, 17 of 20 patients in the TCD group and two of three patients in the non-TCD group achieved complete remission. Donor lymphocyte infusion induced GVHD in nine of 23 patients. Thirty (65%) of 46 patients in the TCD group and 27 (69%) of 39 assessable patients in the non-TCD group remained alive without evidence of disease. The estimated 3-year overall survival rates were similar for the TCD group and the non-TCD group (72% v 68%, respectively; P = .38). At last follow-up, there was no difference in the overall prevalence of GVHD or the proportion of patients requiring immunosuppressive agents between groups. CONCLUSION: These results suggest that the combination of T-cell depletion and post-BMT DLI is a viable treatment option for patients undergoing allogeneic BMT for CML and should be prospectively compared with traditional forms of GVHD prophylaxis.

CD6+ donor marrow T-cell depletion as the sole form of graft-versus-host disease prophylaxis in patients undergoing allogeneic bone marrow transplant from unrelated donors.

PURPOSE: The role of donor marrow T-cell depletion (TCD) in preventing graft-versus-host disease (GVHD) after transplantation of unrelated allogeneic marrow remains undefined. Because different TCD methodologies differ in the degree and specificity with which T cells are removed, it is likely that transplant outcomes would depend on which technique is used. Herein, we report results in the first 48 recipients of unrelated marrow using CD6+ TCD as the sole form of GVHD prophylaxis. PATIENTS AND METHODS: Median age of patients was 46 years (20 to 58 years). Donors were matched at A/B HLA loci. Ablation consisted of cyclophosphamide and fractionated total-body irradiation (TBI; 14 Gy). To facilitate engraftment, patients also received 7.5 Gy (22 patients) [corrected] or 4.5 Gy (26 patients) [corrected] of total lymphoid irradiation (TLI) before admission. No additional immune suppressive prophylaxis was administered. Granulocyte colony-stimulating factor was administered daily from day +1 to engraftment. RESULTS: All 48 patients demonstrated neutrophil engraftment. An absolute neutrophil count of 500 x 10(6)/L was achieved at a median of 12 days (range, 9 to 23 days). There were no cases of late graft failure. The number of CD34+ cells infused/kg was associated with speed of platelet and neutrophil recovery. The dose of TLI did not influence engraftment. Grades 2-4 acute GVHD occurred in 42% of patients (95% confidence interval [CI], 0.28 to 0.57). Mortality at day 100 was 19%. There have been only five relapses. Estimated 2-year survival was 44% (95% CI, 0.28 to 0.59) for the entire group, 58% for patients less than 50 years of age. In multivariable analysis, age less than 50 years (P =.002), cytomegalovirus seronegative status (P =.04), and early disease status at bone marrow transplant (P =.05) were associated with superior survival. CONCLUSION: CD6+ TCD does not impede engraftment of unrelated bone marrow after low-dose TLI, cyclophosphamide, and TBI. CD6+ TCD as the sole form of GVHD prophylaxis results in an incidence of GVHD that compares favorably with many adult studies of unrelated transplantation using unmanipulated marrow and immune-suppressive medications, especially in light of the median age of our patients (46 years). Although event-free survival in patients less than 50 years of age is very encouraging, older patients experience frequent transplantation-related complications despite TCD.

Final analysis of the ECOG I-COPA trial (E6484) in patients with non-Hodgkin's lymphoma treated with interferon alfa (IFN-alpha2a) plus an anthracycline-based induction regimen.

The Eastern Cooperative Oncology Group (ECOG) performed a prospectively randomized study (E6484) evaluating the use of interferon alfa 2a (IFN-alpha2a) in patients with aggressive low-grade or with intermediate-grade non-Hodgkin's lymphoma (NHL) accruing close to 300 patients between 1985 and 1988. Patients were eligible for study if they had bulky or symptomatic low-grade lymphoma or defined intermediate-grade subtypes. Of 291 patients enrolled, 249 were eligible for analysis. All patients were randomized to receive a four-drug cytotoxic chemotherapy regimen including cyclophosphamide, doxorubicin, vincristine and prednisone in 4-week cycles with or without IFN-alpha2a in addition (COPA vs I-COPA). Treatment was given for up to 8-10 months. This report, at a time when the median follow-up among survivors has reached 12 years, updates the analysis of time to treatment failure (TTF), duration of disease-free survival (DFS), and overall survival. Patients randomized to receive IFN-alpha2a had a prolonged TTF (P= 0.008; median 2.4 vs 1.6 years). DFS for those patients who had complete responses was also longer if IFN-alpha2a had been given (P = 0.035; median 2.7 vs 1.8 years). There was a clinically but not a statistically significant prolongation of overall survival by IFN-alpha2a (P= 0.107; median 7.8 vs 5.7 years). There were fewer deaths over time due to lymphoma in patients receiving IFN-alpha2a (67 vs 80 deaths). A subset analysis, based on disease histology (low-grade, follicular, intermediate-grade), revealed a significant prolongation of TTF in patients receiving IFN-alpha2a with either low-grade (P = 0.002; median 2.4 vs 1.6 years) or follicular (P= 0.01; median 2.5 vs 1.7 years) NHL but not intermediate grade (P = 0.622; median 2.3 vs 1.6 years) NHL. This analysis, performed approximately 12 years after closure of the study to accrual, supports the addition of interferon alfa to an induction cytotoxic chemotherapy regimen including cyclophosphamide and doxorubicin in the treatment of follicular NHL.

Phase I/II trial of everolimus in combination with bortezomib and rituximab (RVR) in relapsed/refractory Waldenstrom macroglobulinemia.

We examined the combination of the mammalian target of rapamycin inhibitor everolimus with bortezomib and rituximab in patients with relapsed/refractory Waldenstrom macroglobulinemia (WM) in a phase I/II study. All patients received six cycles of the combination of everolimus/rituximab or everolimus/bortezomib/rituximab followed by maintenance with everolimus until progression. Forty-six patients were treated; 98% received prior rituximab and 57% received prior bortezomib. No dose-limiting toxicities were observed in the phase I. The most common treatment-related toxicities of all grades were fatigue (63%), anemia (54%), leucopenia (52%), neutropenia (48%) and diarrhea (43%). Thirty-six (78%) of the 46 patients received full dose therapy (FDT) of the three drugs. Of these 36, 2 (6%) had complete response (90% confidence interval (CI): 1-16). In all, 32/36 (89%) of patients experienced at least a minimal response (90% CI: 76-96%). The observed partial response or better response rate was 19/36 (53, 90 CI: 38-67%). For the 36 FDT patients, the median progression-free survival was 21 months (95% CI: 12-not estimable). In summary, this study demonstrates that the combination of everolimus, bortezomib and rituximab is well tolerated and achieved 89% response rate even in patients previously treated, making it a possible model of non-chemotherapeutic-based combination therapy in WM.

Induction of replicative senescence by 5-azacytidine: fundamental cell kinetic differences between human diploid fibroblasts and NIH-3T3 cells.

To analyse the putative role of methylation of cytosine residues in the nuclear DNA as a regulatory step during cellular ageing, we incubated ageing human amniotic fluid derived fibroblast-like cells and non-ageing NIH-3T3 cells with 5-azacytidine. BrdUrd/Hoechst and acridine orange (AO) flow cytometry was used to compare the effects of the base analogue on cell proliferation and cell differentiation. In NIH-3T3 cultures, 96h exposures to 4 microM 5-azacytidine caused diminished cell proliferation due to cell arrest in the G1 compartments of the second and third cell cycles of serum stimulated cells. The exit from the G0/G1 compartment was not affected. The 5-azacytidine induced cell kinetic disturbances were unstable in NIH-3T3 cultures, such that pre-treated cells reverted to normal cell cycle transit within 2-3 days after termination of treatment. In contrast, 5-azacytidine pre-treated amniotic fluid derived fibroblast-like cell cultures showed persistently elevated G2 phase arrests and delayed G0/G1 phase exit kinetics, which explain the premature cessation of proliferation observed in these primary cultures. In both cell systems, 5-azacytidine exposed cultures showed elevated numbers of G1 phase cells with increased RNA content as revealed by AO flow cytometry. Again, this effect was reversible in NIH-3T3 cells but not in amniotic fluid derived fibroblast-like cells. These contrasting responses to 5-azacytidine are likely to reflect intrinsic differences in methylation patterns or de novo methylase activity between ageing cell strains and non-ageing cell lines.

Case report of conversion symptom associated with major depressive disorder in a child.

A 10-year-old boy had a prominent conversion symptom that obscured a major depressive episode as defined by DSM-III. After the conversion symptom was removed with amobarbital, his depressive symptoms became more obvious and subsequently improved with imipramine treatment.

Adult psychiatric disorders in psychiatrically ill young adolescents.

Adolescents aged 12-15 years, randomly selected from a psychiatric outpatient clinic, psychiatric consultation service, and general pediatric outpatient clinic, were given a complete psychiatric evaluation and structured diagnostic interview. Of 121 subjects studied, 100 satisfied Rutter and associates' criteria for a childhood psychiatric disorder. When these subjects were rediagnosed according to Feighner and associates' research diagnostic criteria (similar to DSM-III criteria), 52 fulfilled the criteria for an adult disorder. Diagnoses included antisocial personality, hysteria, schizophrenia, depression, mental retardation, anxiety neurosis, and undiagnosed psychiatric illness. There was a correlation between diagnosis according to Rutter and associates' criteria and adult diagnosis.

Comparison of fluorescent polarization immunoassay and radioimmunoassay in measuring cortisol levels in prepubertal depressed children.

Cortisol levels of 21 hospitalized prepubertal depressed children given the dexamethasone suppression test (DST) were measured by radioimmunoassay and by fluorescent polarization immunoassay, a new assay method. Correlation analyses demonstrated a highly significant linear relationship between the two methods of measuring cortisol. Thus, it may be possible to use fluorescent polarization immunoassay to measure cortisol levels in children undergoing the DST.

Psychosocial functioning in children after the death of a parent.

Eight weeks after the death of a parent, children from stable families (N = 38) were compared to depressed inpatients (N = 38) and normal children (N = 19). School behavior, interest in school, peer involvement, peer enjoyment, and self-esteem were similar for bereaved and normal children. Bereaved children functioned significantly better than depressed inpatients. As a group, the bereaved children from stable families did not experience significant, acute psychosocial dysfunction.

Plasma levels of imipramine and adverse effects in children.

In 22 prepubertal depressed children, the total plasma concentration of imipramine and its major metabolite, desipramine, varied by more than sevenfold. The plasma drug concentrations correlated with slowing of intracardiac conduction, elevation of diastolic blood pressure, and increase in heart rate. These drug-induced changes were uniformly observed when the total tricyclic level exceeded 225 ng/ml. However, subjective reporting of nuisance side effects was not related to plasma drug concentration. These findings suggest that children can be safely treated when their plasma levels are below 225 ng/ml. If higher plasma levels are attained, closer monitoring is warranted.

Depression in recently bereaved prepubertal children.

OBJECTIVE: The purpose of this study was to ascertain depressive symptoms in recently bereaved prepubertal children and compare these symptoms with those of depressed prepubertal children. METHOD: The subjects were 38 children who had recently experienced the death of one but not both of their parents. They had to meet strict inclusion criteria so that the effects of bereavement per se, rather than other significant stressors, could be assessed. The comparison group consisted of 38 hospitalized, depressed children individually matched to each bereaved subject for age, sex, and socioeconomic status. All of the children underwent systematic and comprehensive evaluation. They and their parents were independently evaluated by trained interviewers using the parent and child versions of the Diagnostic Interview for Children and Adolescents. Family histories and basic demographic information were also obtained. RESULTS: The recently bereaved children endorsed many depressive symptoms. Thirty-seven percent of them met the DSM-III-R criteria for a major depressive episode. The depressed children, however, had more depressive symptoms on average than the bereaved children. The factors associated with increased depressive symptoms in the bereaved children were 1) the mother as the surviving parent, 2) preexisting untreated psychiatric disorder in the child, 3) family history of depression, and 4) high socioeconomic status. CONCLUSIONS: A considerable number of the bereaved children developed the clinical symptoms of a major depressive episode immediately after the death of a parent. The relation of these symptoms to the subsequent course of grief and to major depressive disorder remains unknown and should be studied further.

Dexamethasone suppression test and clinical outcome in prepubertal depressed children.

Hospitalized prepubertal depressed children with abnormal dexamethasone suppression test (DST) results were treated and given repeat DSTs at 6 weeks (N = 21) and 5 months (N = 14). DST results were significantly correlated with clinical status at 5 months but not at 6 weeks.

Lithium treatment of manic episodes with psychotic features in prepubertal children.

The authors describe the manic symptoms, family psychiatric histories, and psychotic symptoms of 10 prepubertal children 6-12 years old who had a DSM-III diagnosis of manic episode with psychotic features. All of the children improved when treated with lithium alone. Improvement in both manic and psychotic symptoms was noted an average of 11 days after lithium administration was started.

The dexamethasone suppression test in hospitalized prepubertal depressed children.

The dexamethasone suppression test was performed on 20 hospitalized prepubertal children who met DSM-III criteria for major depressive disorder. Fourteen children (70%) did not suppress cortisol secretion at either 8:00 a.m. or 4:00 p.m. The 4:00 p.m. value alone predicted 93% of the nonsuppressors.