Sixteen-week analysis of unaltered elastomeric chain relating in-vitro force degradation with in-vivo extraction space tooth movement.

INTRODUCTION: The purposes of this study were to evaluate whether unaltered elastomeric chain can continue to move teeth for 16 weeks and to relate it to the amount of force remaining for the same batch of elastomeric chains. METHODS: The in-vivo portion of the study had a sample of 30 paired extraction space sites from 22 subjects who were measured for closure of the space every 28 days. The altered side elastomeric chain served as the control and was replaced at 28-day intervals whereas the experimental side remained unaltered. In the in-vitro portion of the study, 100 each of 2-unit and 3-unit segments of the same batch of elastomeric chains were placed in a water bath, and the force was measured for 20 of each segment length at the 28-day measurement points. RESULTS: Statistically significant amounts of space closure occurred at both the altered and unaltered sites at all measurement time points. The mean space closure at the altered sites was minimally greater than that observed at the paired unaltered sites. The mean differences of space closure between the altered and unaltered sites ranged from a minimum of -0.05 mm at 4 weeks to a maximum of -0.14 mm at 8 weeks. The elastomeric chain force degraded rapidly by 4 weeks but continued a gradual diminution of force to 86 g at 16 weeks. CONCLUSIONS: Unaltered elastomeric chain continued to move teeth into extraction spaces for 16 weeks in this sample from both statistically and clinically significant standpoints. There were minimal and statistically insignificant differences in the mean space closure measurements between the paired altered and unaltered sites. The elastomeric chain force at 16 weeks was less than 100 g, yet at the same time point, teeth continued to move clinically.

Alefacept combined with tacrolimus, mycophenolate mofetil and steroids in de novo kidney transplantation: a randomized controlled trial.

Memory T cells play a central role in mediating allograft rejection and are a rational target for immunosuppressive therapy. Alefacept is a recombinant LFA3/IgG1 fusion protein that reduces the number of memory T cells in both psoriatic lesions and the peripheral circulation of psoriasis patients. This study evaluated the efficacy and safety of alefacept compared with placebo when combined with tacrolimus, mycophenolate mofetil and corticosteroids in de novo renal transplant recipients. Between December 2007 and March 2009 patients were randomized in a double-blind fashion to receive alefacept (n = 105) or placebo (n = 107) for 3 months and were then followed for a further 3 months. The primary efficacy endpoint was the incidence of biopsy-confirmed acute T cell mediated rejection (Banff grade ≥ 1) through Month 6. Memory T cell counts were significantly reduced in the alefacept group from Week 3 to study end compared with placebo. However, there was no significant difference between the alefacept and placebo groups for the primary efficacy endpoint (alefacept, 11.0% vs. placebo, 7.0%, p = 0.3). Patient and graft survival as well as renal function was similar between treatment groups. Safety and tolerability were generally similar between the treatment arms. Malignancy was higher in the alefacept treatment arm.

A phase 1, randomized ascending single-dose study of antagonist anti-human CD40 ASKP1240 in healthy subjects.

This first-in-human, phase I study evaluated the safety, tolerability, pharmacokinetic and pharmacodynamic profile of ASKP1240 in healthy subjects. Twelve sequential groups (each 6 active and 3 placebo) were randomly assigned to placebo or single ascending doses of intravenous ASKP1240 (0.00003-10 mg/kg). ASKP1240 exhibited nonlinear pharmacokinetics, with mean maximal serum concentrations and area under the serum concentration-time curves ranging from 0.7 to 251.6 µg/mL and 6.5 to 55409.6 h·µg/mL following doses 0.1 mg/kg-10 mg/kg, respectively. CD40 receptor occupancy by ASKP1240, which was dose-dependent, reached a maximum at doses above 0.01 mg/kg. ASKP1240 was well tolerated, with no evidence of cytokine release syndrome or thromboembolic events. Treatment emergent antibodies to ASKP1240 were detected in 5/70 (7.1%) ASKP1240 recipients. In conclusion, antagonism of the CD40/CD154 interaction with ASKP1240 was safe and well tolerated at the doses tested.

Effect of corticosteroid withdrawal on tacrolimus and mycophenolate mofetil exposure in a randomized multicenter study.

As corticosteroid-sparing protocols are increasingly utilized in kidney transplant recipients, it is crucial to understand potential drug interactions between tacrolimus (TAC) and the effect of corticosteroid withdrawal as well as to characterize dose adjustments of mycophenolate mofetil (MMF) in this setting. This prospective, multicenter, randomized, double-blind study included 397 patients who were randomized on posttransplant day 8 to receive either placebo (CSWD) or corticosteroid continuance (CCS). TAC trough levels at week two posttransplant were significantly greater in the CSWD group whereas TAC doses were comparable to the CCS group. This interaction was not observed in the African American subgroup. Higher serum creatinine and potassium levels were also observed in the CSWD group. MMF dose was significantly reduced in the CSWD group by the investigators because of decreased WBC counts, mostly outside of study protocol criteria, despite similar incidence of neutropenia and reported cytomegalovirus infection. Understanding TAC and MMF exposure in the context of corticosteroid-sparing protocols should allow for improved dosing of immunosuppressants and better management of posttransplant patients.

Modification of the structural stability of human serum albumin in rheumatoid arthritis.

Differential scanning calorimetry (DSC) can indicate changes in structure and/or concentration of the most abundant proteins in a biological sample via heat denaturation curves (HDCs). In blood serum for example, HDC changes result from either concentration changes or altered thermal stabilities for 7-10 proteins and has previously been shown capable of differentiating between sick and healthy human subjects. Here, we compare HDCs and proteomic profiles of 50 patients experiencing joint-inflammatory symptoms, 27 of which were clinically diagnosed with rheumatoid arthritis (RA). The HDC of all 50 subjects appeared significantly different from expected healthy curves, but comparison of additional differences between the RA and the non-RA subjects allowed more specific understanding of RA samples. We used mass spectrometry (MS) to investigate the reasons behind the additional HDC changes observed in RA patients. The HDC differences do not appear to be directly related to differences in the concentrations of abundant serum proteins. Rather, the differences can be attributed to modified thermal stability of some fraction of the human serum albumin (HSA) proteins in the sample. By quantifying differences in the frequency of artificially induced post translational modifications (PTMs), we found that HSA in RA subjects had a much lower surface accessibility, indicating potential ligand or protein binding partners in certain regions that could explain the shift in HSA melting temperature in the RA HDCs. Several low abundance proteins were found to have significant changes in concentration in RA subjects and could be involved in or related to binding of HSA. Certain amino acid sites clusters were found to be less accessible in RA subjects, suggesting changes in HSA structure that may be related to changes in protein-protein interactions. These results all support a change in behavior of HSA which may give insight into mechanisms of RA pathology.

Little salt spring, Florida: a unique underwater site.

Little Salt Spring in southwest Florida, consisting of a shallow, water-filled basin above a deep, vertical underwater cavern, was a freshwater cenote in the peninsula's drier past. It collected and preserved perishable organic artifacts and other evidence of Paleo-Indian and Archaic Indian origin ranging in age from 12,000 to 9000 and from 6800 to 5200 years ago. An Archaic Period cemetery containing an estimated 1000 burials occupies an adjoining muck-filled slough and presently drowned portions of the basin of the spring. Artifacts and the nature of interment suggest a cultural link between the Archaic people and the much later Glades Tradition of southern Florida.

Development of a continuous direct compression platform for low-dose drug products.

In this work a continuous direct compression process was developed for a low-dosed drug product. Each unit operation of the GEA CDC-50 system was thoroughly investigated. This paper aimed to tackle the macroscopic and microscopic blend uniformity challenges inherently associated with continuous direct compression of cohesive and agglomerated APIs formulated at low dose. Density, compressibility and flow were identified as key material properties at the feeding stage. The screw speed coupled with powder flow regulated the gravimetric feeding performance. The impact of process and design variables was elucidated at the blending stage. The impeller configuration (number and pattern of radial mixing blades) and speed were key variables to steer the residence time distribution at the blending stage. An impeller configuration with distributed radial mixing blades could sufficiently filter the steady state feeding variability at low mixer speed, but exerted limited strain and shear on the blend. Hence micro-agglomerates persisted through the blending process and occasionally resulted in super potent tablets. Therefore, a new configuration was evaluated with more radial mixing blades centered on the impeller. This configuration resulted in a long mixing time at high tip speed which induced a maximized strain and shear. Consequently, excellent uniformity of the blend and tablets at macroscopic and microscopic level was achieved. Besides, this impeller improved robustness towards feeding disturbances, changes in process settings and variable blend properties. Next, it was demonstrated that the lubrication step requires critical attention during the design of the equipment, formulation and process. This study provided abundant evidence that an optimized continuous direct compression process allows direct compression of challenging low-dose drug products.

Continuous direct compression as manufacturing platform for sustained release tablets.

This study presents a framework for process and product development on a continuous direct compression manufacturing platform. A challenging sustained release formulation with high content of a poorly flowing low density drug was selected. Two HPMC grades were evaluated as matrix former: standard Methocel CR and directly compressible Methocel DC2. The feeding behavior of each formulation component was investigated by deriving feed factor profiles. The maximum feed factor was used to estimate the drive command and depended strongly upon the density of the material. Furthermore, the shape of the feed factor profile allowed definition of a customized refill regime for each material. Inline NIRs was used to estimate the residence time distribution (RTD) in the mixer and monitor blend uniformity. Tablet content and weight variability were determined as additional measures of mixing performance. For Methocel CR, the best axial mixing (i.e. feeder fluctuation dampening) was achieved when an impeller with high number of radial mixing blades operated at low speed. However, the variability in tablet weight and content uniformity deteriorated under this condition. One can therefore conclude that balancing axial mixing with tablet quality is critical for Methocel CR. However, reformulating with the direct compressible Methocel DC2 as matrix former improved tablet quality vastly. Furthermore, both process and product were significantly more robust to changes in process and design variables. This observation underpins the importance of flowability during continuous blending and die-filling. At the compaction stage, blends with Methocel CR showed better tabletability driven by a higher compressibility as the smaller CR particles have a higher bonding area. However, tablets of similar strength were achieved using Methocel DC2 by targeting equal porosity. Compaction pressure impacted tablet properties and dissolution. Hence controlling thickness during continuous manufacturing of sustained release tablets was crucial to ensure reproducible dissolution.

Hepatocyte injury during post-operative sepsis: activated neutrophils as potential mediators.

We evaluated the concept that the vascular entrance of both bacterial and nonbacterial particulate material could lead to hepatic parenchymal cell injury, either due to postphagocytic Kupffer cell activity or the margination of activated leukocytes in the liver. Injection of denatured, collagen-coated particles as well as heat-killed bacteria were used as particulate challenges. Hepatic parenchymal cell injury in vivo during postoperative sepsis was evaluated by plasma aspartate aminotransferase (AST) and ornithine carbamyl transferase (OCT) enzyme levels over 3-72 h. AST and OCT levels elevated following either laparotomy plus cecal ligation (mild sepsis) or laparotomy plus cecal ligation with puncture (severe sepsis), with the peak level at 24 h. In addition, the direct intravenous injection of either nonbacterial foreign particles or heat-killed Pseudomonas aeruginosa into normal rats also produced a dose-dependent elevation of AST and OCT. The plasma level of either AST or OCT actually increased 350-400% after injection of the non-bacterial particles. A similar dose related elevation in enzymes followed the intravenous injection of heat-killed Pseudomonas. To differentiate the potential contribution of activated hepatic Kupffer cells versus activated marginated neutrophils to the in vivo hepatic injury, we determined the release of the hepatic specific enzyme OCT by cultured hepatic parenchymal cells when they were exposed to isolated Kupffer cells or isolated PMNs that were activated by exposure to dead bacteria. Bacteria alone when added to cultured hepatocytes did not induce significant OCT release. In contrast, activated PMNs but not Kupffer cells induced a significant (p less than 0.05) release of OCT from parenchymal cells into the culture media. Thus, in vivo transient hepatic parenchymal cell injury with post-operative sepsis may be mediated by the margination of activated PMNs in the liver.

Draft Genome Sequence of Erwinia billingiae OSU19-1, Isolated from a Pear Tree Canker.

Plant-associated Erwinia include pathogenic and nonpathogenic species. We report the 5.6-Mb genome sequence of Erwinia billingiae OSU19-1, isolated from a canker on a pear tree inoculated with Erwinia amylovora. OSU19-1 and a closely related European isolate, E. billingiae Eb661(T), share many similarities including 40 kb of plasmid sequence.

Estrogen and progesterone production by granulosa cell monolayers derived from in vitro fertilization procedures: lack of evidence for modulation by androgen.

The role(s) of androgens in the steroidogenic regulation of human granulosa cell production of estrogen and progesterone during monolayer culture was studied. These cells were exposed in vivo to human menopausal gonadotropin and hCG gonadotropin with or without clomiphene citrate. Steroid production rates were compared between cells cultured in control medium and those cultured in medium containing a nonaromatizable androgen [dihydrotestosterone (DHT)] or an aromatizable androgen [androstenedione (A'D)]. Some cultures received A'D from 3-12 days; other cultures received DHT alone for 3, 6, or 9 days before the addition of A'D for 3 days. The effect on steroid production during the culture interval before the addition of A'D also was evaluated. Exposure to A'D increased estrogen production over 50-fold compared with that in control cells or those treated with DHT (P less than 0.001). DHT also failed to alter estrogen production when A'D was added to cultures. Furthermore, the delay in introducing A'D to the cultures for up to 9 days did not decrease subsequent estrogen production compared with that in cultures continually exposed to A'D or DHT plus A'D. Progesterone production was substantial for at least 12 days of culture and was unaffected by the presence of androgen. These results do not confirm previous studies using murine or porcine granulosa cells, which suggested that androgen receptor-dependent mechanisms were involved in increasing estrogen and/or progesterone production in vitro. Rather, they indicate that androgen may not be required to maintain aromatase capability per se in human granulosa-luteal cells previously exposed to ovulation-inducing quantities of gonadotropin.

Expression and function of the human calcitonin/alpha-CGRP gene in health and disease.

Studies on the structure and expression of the rat and human calcitonin gene provide a remarkable example by which a combination of molecular, immunocytochemical and pharmacological techniques have led to the identification of a novel gene product, the calcitonin gene-related peptide, a peptide of unexpected tissue distribution and biological activity. Future studies at the molecular level will provide insight into post-transcriptional mechanisms by which a single gene can give rise to discrete mRNAs in a tissue-specific manner. The isolation and characterization of the CGRP receptor, and hence the site(s) and mechanism(s) of action of the CGRP family within the vasculature, will also add significantly to our understanding of molecular mechanisms involved in the modulation of cardiovascular function in man. Furthermore, the role and mechanism of action of the CGRP peptide family in the central and peripheral nervous systems remains to be elucidated. It is also apparent that measurement of circulating plasma levels of CGRP may be of diagnostic and prognostic value, providing information concerning the onset and progression of lung and thyroid carcinoma. Our analysis of the structure and expression of the calcitonin/alpha-CGRP gene also demonstrates, for the first time, the molecular basis of large calcitonin secretion by lung carcinoma cells. Whether the expression of this gene in lung carcinoma cell-lines can truly be described as 'ectopic' is however questionable, in the light of recent immunocytochemical evidence which demonstrates that the lung is a major source of CGRP producing cells in the rat (see Springall et al., 1984).

Reversal of opsonic deficiency in surgical, trauma, and burn patients by infusion of purified human plasma fibronectin. Correlation with experimental observations.

Plasma fibronectin deficiency has been documented in critically ill surgical, trauma, and burn patients. Human plasma fibronectin was isolated by gelatin-Sepharose affinity chromatography and evaluated with respect to its opsonic activity following pasteurization, its in vivo clearance kinetics, and its short-term influence on cardiovascular hemodynamics in postoperative septic sheep. Six patients with low plasma fibronectin levels were also evaluated with respect to temporal changes of immunoreactive fibronectin and opsonic activity following infusion of fibronectin at a dose calculated to elevate the plasma fibronectin level to 400 micrograms/ml. With utilization of three different in vitro radioisotopic phagocytic assays, i.e., liver slice assay, peritoneal macrophage monolayer assay, and Kupffer cell monolayer assay, retention of opsonic activity by fibronectin following pasteurization was documented. The normal biphasic kinetics associated with plasma clearance of fibronectin were also not altered by pasteurization. In postoperative septic sheep with hemodynamic instability, intravenous infusion of 500 mg of purified human fibronectin initiated no abnormal hemodynamic response. Indeed, as compared with placebo, the infusion of fibronectin into the postoperative septic sheep resulted in a more stable systemic vascular resistance and pulmonary vascular resistance with a higher arterial pressure. It also elevated immunoreactive fibronectin levels (p less than 0.05) and increased opsonic activity (p less than 0.05). Surgical, trauma, and burn patients (ages 18 to 80 years) with low plasma fibronectin levels (160 to 236 micrograms/ml) manifested no disturbance in cardiovascular, respiratory, or hematologic parameters following fibronectin infusion (590 to 988 mg per patient), but did display an early increase of opsonic activity. This standardized, pasteurized, and opsonically active preparation of purified human plasma fibronectin (5.0 mg/ml after reconstitution) has utility for future randomized clinical trials in injured patients with sepsis.

The effects of oral pentoxifylline on the cytokine release syndrome during inductive OKT3.

The cytokine release syndrome (CRS) accompanying OKT3 therapy is a major cause of posttransplant morbidity. The pathogenesis of this syndrome has been attributed to the synthesis of tumor necrosis factor, interleukin 2 (IL-2), interleukin 6 (IL-6), and gamma-interferon in response on T lymphocyte stimulation by OKT3. The hemorrheologic agent pentoxifylline (PTX) inhibits the synthesis of TNF alpha in vitro in response to a variety of stimuli, including OKT3. We performed a randomized, double-blinded trial of PTX during OKT3 induction in recipients of cadaveric renal allografts. Patients received either PTX 800 mg or placebo 2 hr before the initial dose of OKT3 and every 8 hr thereafter during the first 3 posttransplant days. Serum TNF alpha and IL-6 concentrations were measured pre-OKT3 and at 2 and 6 hr post-OKT3 on the first 3 posttransplant days. Despite the achievement of apparently adequate plasma levels of PTX and its active metabolites, no difference was observed in the incidence or severity of clinical manifestations of CRS. Serious manifestations of CRS--including acute pulmonary edema, encephalopathy, and aseptic meningitis--were not seen in either group. Serum TNF alpha and IL-6 concentrations were similar in PTX and control patients throughout the course of the study. Plasma levels of PTX and its active metabolites did not correlate with serum TNF alpha levels, serum IL-6 levels, or the incidence and severity of clinical manifestations of CRS.

Fibronectin and the critically ill patient: current status.

In summary, deficiency of plasma fibronectin has now been documented in a variety of clinical entities. Persistently low fibronectin may have prognostic value, and in certain patients may provide a clue to occult sepsis and potential organ failure. The clinical benefit of infusion of fibronectin-rich cryoprecipitate or purified human plasma fibronectin has yet to be determined in well-controlled randomized clinical trials. However, if such results become available then infusion of plasma fibronectin may provide a valuable therapeutic modality in the care of the critically-ill patient.

Human alpha- and beta-CGRP and rat alpha-CGRP are coronary vasodilators in the rat.

The effects of the recently described human alpha-calcitonin gene-related peptide (CGRP), human beta-CGRP and rat alpha-CGRP have been compared with those of the vasodilator sodium nitroprusside, on the rat and rabbit isolated heart. Hearts were perfused at constant flow and [Arg8]-vasopressin was used to increase coronary perfusion pressure. In the rat heart, the order of potency for evoking cumulative dose-dependent falls in perfusion pressure was human beta-CGRP greater than rat alpha-CGRP greater than human alpha-CGRP greater than sodium nitroprusside. In the same preparations the three CGRPs (but not sodium nitroprusside) elicited cumulative dose-related increases in heart rate. In the rabbit heart the order of potency for vasodilatation was rat alpha-CGRP greater than human alpha-CGRP greater than sodium nitroprusside. In marked contrast to results from the rat, neither rat alpha-CGRP nor human alpha-CGRP altered heart rate in the rabbit isolated heart. These results show that human alpha- and beta-CGRP and rat alpha-CGRP are vasodilators in the coronary vasculature, but that there is species variation as CGRP had a positive chronotropic effect in the rat heart but not in the rabbit heart.

Human and rat alpha-CGRP but not calcitonin cause mesenteric vasodilatation in rats.

A single gene encodes both calcitonin and the calcitonin gene-related peptide (CGRP). Human and rat alpha-CGRP were compared with sodium nitroprusside in the rat and rabbit isolated mesenteric vascular preparation perfused at constant flow. In the presence of the vasoconstrictor noradrenaline (10(-5) M), rat alpha-CGRP was about 10 times as potent as either human alpha-CGRP or sodium nitroprusside as a vasodilator in the rat mesenteric vasculature. In the rabbit mesenteric vasculature the order of potency was rat alpha-CGRP greater than human alpha-CGRP greater than sodium nitroprusside. Human and salmon calcitonin showed no vasodilator activity at doses 100 times greater than human alpha-CGRP. These results show that human and rat alpha-CGRP are potent vasodilators in the mesenteric vasculature, an effect not mimicked by the alternative gene product, the plasma calcium lowering hormone calcitonin.

Type of hemodialysis and preference for behavioral involvement: interactive effects on adherence in end-stage renal disease.

Examined the effects of hemodialysis type (i.e., staff controlled, in center vs. patient controlled, home) and patient preference for behavioral involvement on adherence and emotional adjustment in a sample of 53 patients with end-stage renal disease. Consistent with person x treatment interaction models, higher levels of preference for behavioral involvement were associated with better dietary adherence (i.e., lower serum potassium) for patients receiving dialysis at home but worse dietary adherence for patients receiving treatment in a dialysis center. A similar though weaker patient x treatment type matching pattern was observed for fluid-intake adherence (i.e., interdialytic weight gain). No effects were observed for patients' self-reported depression levels. Possible mechanisms for the interactional effect on adherence are discussed.