The temporal expression of estrogen receptor alpha-36 and runx2 in human bone marrow derived stromal cells during osteogenesis.

During bone maintenance in vivo, estrogen signals through estrogen receptor (ER)-α. The objectives of this study were to investigate the temporal expression of ERα36 and ascertain its functional relevance during osteogenesis in human bone marrow derived stromal cells (BMSC). This was assessed in relation to runt-related transcription factor-2 (runx2), a main modulatory protein involved in bone formation. ERα36 and runx2 subcellular localisation was assessed using immunocytochemistry, and their mRNA expression levels by real time PCR throughout the process of osteogenesis. The osteogenically induced BMSCs demonstrated a rise in ERα36 mRNA during proliferation followed by a decline in expression at day 10, which represents a change in dynamics within the culture between the proliferative stage and the differentiative stage. The mRNA expression profile of runx2 mirrored that of ERα36 and showed a degree subcellular co-localisation with ERα36. This study suggests that ERα36 is involved in the process of osteogenesis in BMSCs, which has implications in estrogen deficient environments.

Altered leucocyte progenitor profile in human bone marrow from patients with major trauma during the recovery phase.

BACKGROUND: Changes in human bone marrow associated with the systemic inflammatory response to injury are little understood. It was hypothesized that major trauma results in an altered bone marrow leucocyte progenitor profile, with either uniform depletion or the balance between multipotent and committed progenitors varying, depending on whether self-renewal is favoured over differentiation. METHODS: Bone marrow aspirate and peripheral blood samples were obtained at definitive surgery in adults with pelvic fractures from blunt trauma (major trauma with Injury Severity Score (ISS) at least 18, or isolated fractures) and control patients undergoing iliac crest bone grafting. ISS, interval to surgery and transfusion in the first 24 h were recorded. Bone marrow aspirate flow cytometry was used to identify haemopoietic progenitor cells (CD34(+) ), multipotent cells (CD34(+) CD45(+) CD38(-) ) and oligopotent cells (CD34(+) CD45(+) CD38(lo/+) and CD34(+) CD45(+) CD38(BRIGHT(++ +)) subsets). Peripheral blood levels of inflammatory markers were measured, and the ratio of immature to mature (CD35(-) /CD35(+) ) granulocytes was determined. RESULTS: The median (range) interval between injury and sampling was 7 (1-21) and 5 (1-21) days in the major trauma and isolated fracture groups respectively. The CD34(+) pool was significantly depleted in the major trauma group (P = 0·017), particularly the CD34(+) CD45(+) CD38(BRIGHT(++ +)) oligopotent pool (P = 0·003). Immature CD35(-) granulocytes increased in bone marrow with increasing injury severity (P = 0·024) and massive transfusion (P = 0·019), and in peripheral blood with increasing interval to surgery (P = 0·005). CONCLUSION: Major blunt trauma resulted in changes in the bone marrow CD34(+) progenitor pool. At the point in recovery when these samples were obtained, oligopotent progenitors were lost from the bone marrow, with continued release of immature cells.

Combing Transcriptomes for Secrets of Deep-Sea Survival: Environmental Diversity Drives Patterns of Protein Evolution.

Ctenophores, also known as comb jellies, live across extremely broad ranges of temperature and hydrostatic pressure in the ocean. Because various ctenophore lineages adapted independently to similar environmental conditions, Phylum Ctenophora is an ideal system for the study of protein adaptation to extreme environments in a comparative framework. We present such a study here, using a phylogenetically-informed method to compare sequences of four essential metabolic enzymes across gradients of habitat depth and temperature. This method predicts convergent adaptation to these environmental parameters at the amino acid level, providing a novel view of protein adaptation to extreme environments and demonstrating the power and relevance of phylogenetic comparison applied to multi-species transcriptomic datasets from early-diverging metazoa. Across all four enzymes analyzed, 46 amino acid sites were associated with depth-adaptation, 59 with temperature-adaptation, and 56 with both. Sites predicted to be depth- and temperature-adaptive occurred consistently near Rossmann fold cofactor binding motifs and disproportionately in solvent-exposed regions of the protein. These results suggest that the hydrophobic effect and ligand binding may mediate efficient enzyme function at different hydrostatic pressures and temperatures. Using predicted adaptive site maps, such mechanistic hypotheses can now be tested via mutagenesis.

Osteogenic Potential of Human Umbilical Cord Mesenchymal Stem Cells on Coralline Hydroxyapatite/Calcium Carbonate Microparticles.

Coralline hydroxyapatite/calcium carbonate (CHACC) is a biodegradable and osteoconductive bone graft material with promising clinical performance. CHACC has been shown to support proliferation and osteogenic differentiation of human bone marrow mesenchymal stem cells (MSCs) in vitro and demonstrated to work as a functional scaffold for bone formation in vivo. Umbilical cord matrix is a more accessible and abundant tissue source of MSCs, but its osteogenic capacity in comparison to human bone marrow when cultured on CHACC has not yet been demonstrated. In this study, we assessed the osteogenic differentiation capacity of human MSCs, isolated from bone marrow and umbilical cord matrix and characterised by flow cytometry, when cultured on 200-300 µm CHACC granules. The 3D cultures were characterised by brightfield and scanning electron microscopy (SEM). Osteogenic potential was assessed by immunocytochemistry and qPCR for key markers of bone differentiation (alkaline phosphatase, runx2, type I collagen, and osteocalcin). By day 1, the MSCs had enveloped the surface of the CHACC granules to form organoids, and by day 7, cells had proliferated to bridge nearby organoids. Extracellular matrix deposition and osteogenic differentiation were demonstrated by MSCs from both tissue sources at day 21. However, MSCs from bone marrow demonstrated superior osteogenic differentiation capability compared to those from umbilical cord matrix. In conclusion, it is possible to culture and induce osteogenic differentiation of umbilical cord matrix MSCs on CHACC. Further research is required to optimise the osteogenicity of umbilical cord matrix MSCs to release their full potential as a readily available, accessible, and abundant tissue source for bone tissue engineering.

Capsaicin desensitization to plasma extravasation evoked by antidromic C-fiber stimulation is not associated with antinociception in the rat.

The relationship of capsaicin desensitization to the antinociception produced by acute capsaicin administration was studied in the rat. Cutaneous application of 1% capsaicin (3 x/day) for 1 day antagonized plasma extravasation evoked by either 5% capsaicin applied to the dorsal surface of the hindpaw or antidromic stimulation of the saphenous nerve. Treatment with either 1% or 5% capsaicin (3 x/day) for up to 4 days failed, however, to produce thermal antinociception as measured by the rat paw-withdrawal procedure. In contrast, intracutaneous injection of 100 micrograms capsaicin produced marked increases in paw-withdrawal response latencies for up to six hours after injection. These data demonstrate a dissociation between block of cutaneous plasma extravasation and thermal antinociception in the rat. The data provide in vivo evidence that desensitization of peripheral nociceptors by capsaicin cannot account for the antinociception observed following acute capsaicin administration. Finally, these data support the notion that some pharmacologic actions of capsaicin in the rat can be dissociated from frank neurotoxicity on the basis of concentration or dose.

Percutaneous absorption of vanilloids: in vivo and in vitro studies.

The percutaneous absorption of three highly lipophilic analogs of capsaicin--vanillylnonanamide (VN), olvanil, and NE-21610--was measured in vivo in the CD:VAF rat, and in vitro through excised CD: VAF and SkH:Fz rat skin and human cadaver skin. Absorption and skin metabolism were monitored by radiolabel techniques. The rank order of penetration in all species was VN > olvanil > NE-21610, in accordance with that expected from their physical properties. Rat skin was more permeable than human skin by factors ranging from 4 to 8 for VN, 10 to 20 for olvanil, and approximately 10 to 100 for NE-21610. All three compounds were extensively metabolized during passage through fresh SkH:Fz rat skin, with the primary route of degradation for at least two of the compounds involving hydrolysis of the amide bond (the metabolites of NE-21610 were not identified). For the in vitro studies a range of receptor solutions was employed to determine a set of conditions that best mimicked in vivo absorption. The results with phosphate-buffered saline containing a preservative and 1-6% polyoxyethylene-20 oleyl ether (Oleth-20) were in good agreement with in vivo results for all three compounds for periods up to 24 h post-dose; after this time, in vivo absorption rates declined but in vitro rates remained relatively constant. Buffered saline or saline containing 0.5% bovine serum albumin led to marked underestimates of in vivo penetration for olvanil and NE-21610, whereas a 1:1 ethanol: water solution led to gross overestimates of the in vivo absorption rates for all three compounds.

Treatment of cervical kyphosis in children.

Children with severe cervical kyphosis present a difficult treatment challenge. The most common etiology of this deformity is extensive laminectomies, especially associated with postlaminectomy irradiation. The deformity can be rapidly progressive leading to neurologic involvement. With intact posterior elements, kyphosis can occur as a result of congenital, traumatic, metabolic or neoplastic processes. Treatment is directed towards early recognition, arrest of the progression of deformity, and improvement of neurologic symptoms. Patients with loss of posterior elements can be treated effectively by preoperative traction and a single-staged anterior release with strut fusion. Patients with intact posterior elements require preoperative traction, initial posterior osteotomies with intraoperative traction, then an anterior release with strut fusion. All patients need rigid postoperative halo immobilization for a minimum of 3 to 4 months to maintain position. Using these techniques, nine patients were treated surgically with satisfactory outcomes.

Effect of dietary carbohydrate type and content on the response of male rats to dietary sodium saccharin.

The role of dietary carbohydrate composition and concentration in the response of male rats to sodium saccharin (NaS) was ascertained by comparing the response to 5% dietary NaS in rats given diets containing 65% starch, 50% sucrose together with 15% starch, 65% glucose, or 3% sucrose. NaS induced similar levels of caecal enlargement and increases in urine volume and bladder mass when given with any of the three forms of carbohydrate at 65% in the diet. However with the 3% sucrose diet, NaS caused a lesser caecal enlargement and no increase in urine volume or bladder mass. These findings suggest that NaS not only inhibits saccharide hydrolysis but also inhibits glucose transport. The significance of these findings in relation to NaS-associated bladder tumours is discussed.

Effect of N-nitroso-n-butyl-(4-hydroxybutyl)amine exposure on the changes in mineral disposition caused by trisodium nitrilotriacetate.

Male Fischer 344 rats were used to determine effect of consumption of 0.5% N-nitroso-n-butyl-(4-hydroxybutyl)amine (BNN) in the drinking-water for 2 wk on the response to 0.02, 0.2 and 2.0% trisodium nitrilotriacetate (Na3 NTA . H2O) in the diet in terms of urinary mineral excretion, bladder mass and bladder mineral concentrations. The primary objective of the study was to determine whether exposure of rats to an initiating dose of a bladder carcinogen (BBN) alters the threshold dose of Na3NTA . H2O required to alter urinary or bladder mineral concentrations or the dose-response to NTA. Such alterations are considered to be necessary precursors for changes in bladder morphology in rats fed NTA in chronic toxicity studies (Anderson, Bishop & Campbell, CRC Crit. Rev. Toxicol. 1985, 15, 1). The results demonstrated that BBN exposure caused an increase in bladder mass and bladder-tissue Zn concentration. However, BBN pretreatment did not have any effect on Na3NTA . H2O metabolism, the threshold dose of Na3NTA . H2O required to attain the necessary conditions for induction of bladder toxicity by NTA, or the dose-response relationships for NTA's effects on any parameter examined. From these data, it is concluded that it is unlikely that NTA would show a different threshold or dose-response for bladder tumour promotion than for its tumorigenicity at this site, which has been demonstrated previously (National Cancer Institute, DHEW Publication No. (NIH) 77-806, 1977).

Urinary and bladder responses to immobilization in male rats.

Immobilization of groups of five to nine male rats for 2-5 days results in a 50% increase in urinary bladder fresh weight compared with normally caged controls. The increase in urinary bladder weight was not due to tissue oedema and was accompanied by epithelial hyperplasia in some urinary bladders. Immobilization did not alter total urine volume, but it did decrease the frequency of urine voiding and doubled the mean urine weight/voiding. Thus, bladder distention caused by the increased volume per voiding caused a rapidly induced increase in bladder tissue growth, and was accompanied by an increase in bladder epithelial cell division.

Renal pelvic and ureteral dilatation in male rats ingesting trisodium nitrilotriacetate.

Renal pelvic and proximal ureteral dilatation has been observed in male Charles River (CD) and Fischer 344 rats ingesting comparable doses of Na3NTA X H2O. Ureteral dilatation is accompanied by epithelial surface ulceration and erosion. Taken together with previous work (Anderson, Alden & Merski, Fd Chem. Toxic. 1982, 20, 105), these findings demonstrate that the ingestion of high doses of NTA results in similar effects on the transitional epithelium throughout the urinary tract.

Traumatic spondylolisthesis of the axis.

A series of 123 patients suffering traumatic spondylolisthesis of the axis is reported. This lesion is associated with extension and axial loading injury, and there is a high incidence of injuries of the face or scalp and of associated fractures of the upper cervical spine. There is a low incidence of neurological injury, which seems paradoxical in the presence of what appears to be gross instability, but protection from extremes of flexion and extension may be adequate treatment. Union is usual regardless of displacement. Traction is a safe means of treatment, but early waking in a halo support reduces time in hospital without jeopardizing the result. Operation is needed only for chronic instability with or without pain, and anterior interbody fusion of C2--3 is than preferred in order to preserve rotation at the atlanto-axial joints.

Traumatic spondylolisthesis of the axis.

The lesions of C2 seen in hyperextension injuries of the cervical spine following motor vehicle accidents, diving accidents, and headlong falls resemble the cervical lesion found in judicial hangings. Although the mechanism of injury in these cervical fractures is different, the distinction is significant. The fracture seen in motor vehicular accidents today usually seems to be one of hyperextension and axial compression rather than hyperextension and distraction. This distinction is responsible for the low incidence of neurologic involvement seen in the fracture caused by motor vehicle accidents. The incidence of face and scalp injuries associated with axis pedicle fractures appears significant. Union of this fracture can generally be expected, and it rarely produces late sequelae. Management of this fracture is generally preferable on an ambulatory basis utilizing a cervicothoracic brace or a halo and caudal support in more unstable fractures. A period of 12 weeks of immobilization is recommended for satisfactory union of this fracture. Occasionally the treatment of this fracture may require traction or operative intervention. If operative intervention is required, an anterior route employing interbody fusion is suggested.

Atlantoaxial rotary deformities.

Persistent torticollis in younger patients, particularly after trivial trauma or an upper respiratory tract infection, suggests a diagnosis of atlantoaxial rotary fixation. The diagnosis can be confirmed by cineroentgenography. Anterior displacement of the atlas, indicating a deficient transverse ligament, should be ruled out by flexion-extension lateral roentgenograms. If conservative management fails to achieve reduction or is followed by a recurrence of the deformity, arthrodesis is indicated.

Atlantal fracture in a child through congenital anterior and posterior arch defects. A case report.

A five-year-old boy sustained a vertical compression injury to the occiput, initial findings were consistent with a cervical spine injury. Evaluation, including plain films, tomograms, and CT scanning, confirmed a C1 fracture through congenital anterior and posterior arch defects. Flexion and extension tomograms showed this rare lesion to be stable, and no surgical intervention was required. Because congenital anomalies of the cervical spine can be confused with traumatic lesions, thorough evaluation is warranted. Treatment should be based on signs of instability, if present.

MAC regulations: their predicted economic impact on pharmaceutical reimbursement.

The Maximum Allowable Cost (MAC) regulations, Part 19 of Subtitle A of Title 45 of the Code of Federal Regulations, were promulgated as a method of cost containment applicable to a component of the escalating health care expenditures. This study predicted the economic impact of the MAC regulations on the Medicaid program of Georgia. Reimbursements for Medicaid prescription drug products by the Georgia Department of Medical Assistance for 1976-1977 were compared with the reimbursements for drug products that would have been paid under the MAC regulation limitations had the MAC regulations been in force during the time period studied. The cost per unit of a purposive sample of 386 drug entities sorted into eight code groups was compared with the MAC-regulated cost per unit reimbursements. Significant differences in cost were noted for three of the eight code groups. For these groups, reimbursements of before MAC-regulated drug product cost per unit were greater than reimbursements for predicted MAC-regulated drug product cost per unit. Further analysis of the data predicted a reduction of reimbursements for the eight classes of prescription drug products studied to be 24.42%. This reduction compares favorably with the projected savings of 22-36% reported by the Task Force on Prescription Drugs.

Osteoid osteoma of the thoracic spine: report of a case.

Osteoid osteoma occurring in the thoracic spine, manifest as purely referred pain, is extremely uncommon. In the case of a 24-year-old man, the treatment for painful osteoid osteoma of the spine was excision. Since excision of a thoracic spinal osteoid osteoma compromised stability, a posterior fusion was performed at the time of excision, and the results were excellent.

Physiological model for tissue glutathione depletion and increased resynthesis after ethylene dichloride exposure.

Ethylene dichloride (EDC) is metabolized by two competing pathways both of which consume glutathione (GSH). EDC undergoes oxidation to form chloroacetaldehyde (CAA) which is detoxified by GSH and also reacts directly with GSH to form 2-(s-chloroethyl)-GSH. A physiological pharmacokinetic model developed for EDC was extended to describe tissue GSH turnover and its depletion after EDC exposures. This GSH model was necessary to keep track of GSH concentrations with time, as EDC metabolism is affected by GSH status. Reactions of GSH with EDC and GSH with CAA were defined as second-order. Steady-state GSH formation was modeled as zero-order and GSH loss as first-order. GSH rebound effects after its depletion were controlled by a GSH synthetase reaction, which allowed time- and GSH concentration-dependent feedback for increased GSH resynthesis. The model was developed for liver GSH in the rat and was extrapolated to include the lung. Allometric scaling was used to extrapolate the model to other animal species. Experimental observations in the rat and mouse were consistent with model predictions.

Trypsin inhibitor ingestion-induced urinary indican excretion and pancreatic acinar cell hypertrophy.

Sodium saccharin (NaSacc) has been shown to be a protease inhibitor and to induce an increase in urinary indican, which is a product that is dependent on microbial metabolism of tryptophan. These findings suggest that urinary indican might provide a noninvasive marker of increased pancreatic acinar cell size associated with plant trypsin inhibitor ingestion. The results demonstrate the 7.5% of dietary NaSacc, which increases urinary indican, also increases relative pancreas mass (g/kg body weight), and that these effects are not induced by intravenous infusion of NaSacc. Dietary soybean trypsin inhibitor in the dose range of 17-713 mg/100 g diet was associated with parallel dose-dependent increases in urinary indican and pancreatic acinar cell size (assessed histologically). These findings suggest that measurement of relative urinary indican excretion (microgram/g diet ingested) can provide a noninvasive marker of increased pancreatic acinar cell size in rats that ingest compounds which inhibit digestive proteases.