Conventional Respiratory Management of Spinal Cord Injury.

Respiratory complications often result from acute spinal cord injury. Ventilatory assistance/support is often required 12 hours to 6 days after admission and is typically delivered via translaryngeal tubes. When not weanable from ventilatory support, tracheostomy tubes are placed. Supplemental O2 is often provided irrespective of whether or not the patient is hypoxic. This renders the oximeter ineffective as a gauge of alveolar ventilation, airway secretion management, and residual lung disease, and can exacerbate hypercapnia. Thus, hypoventilation and airway secretions must be effectively treated to prevent lung disease and to maintain normal O2 saturation and CO2 levels without supplemental O2.

Noninvasive Respiratory Management of Spinal Cord Injury.

Intubated ventilator-dependent patients with high-level spinal cord injury can be managed without tracheostomy tubes provided that they have sufficient cognition to cooperate and that any required surgical procedures are completed and they are medically stable. Intubation for a month or more than extubation to continuous noninvasive ventilatory support (NVS) can be safer long term than resort to tracheotomy. Noninvasive ventilation (NIV) is not conventionally being used for ventilatory support. Noninvasive interfaces include mouthpieces, nasal and oronasal interfaces, and intermittent abdominal pressure ventilators. NIV/NVS should never been used without consideration of mechanical insufflation-exsufflation for airway secretion clearance.

A Mechanical Intermittent Abdominal Pressure Ventilator.

Pneumatically driven intermittent abdominal pressure ventilators were a popular means of daytime ventilatory support until the late 1960s paradigm shift to invasive (tracheostomy) mechanical ventilation. However, although many patients still use intermittent abdominal pressure ventilators, currently available turbine-driven portable home care ventilators are not powerful enough to always successfully operate them. We describe a new mechanically driven intermittent abdominal pressure ventilator operated by a 1-pound motor that provided a depth of abdominal compression of almost 2 in in 1.05 to 1.13 secs to normalize alveolar ventilation for a 72-yr-old postpolio survivor. It increased her autonomous 200- to 320-ml tidal volumes by greater than 300 ml to normalize her respiratory rate, relieve her dyspnea, and maintain normal oxyhemoglobin saturation levels throughout daytime hours for a 9-mo period of continuous ventilatory support.

Mechanical In-exsufflation-Expiratory Flows as Indication for Tracheostomy Tube Decannulation: Case Studies.

Mechanical insufflation exsufflation-expiratory flows (MIE-EFs) correlate with upper airway patency. Patients dependent on continuous noninvasive ventilatory support with severe spinal muscular atrophy type 1, now over 20 yrs old, have used MIE sufficiently effectively along with continuous noninvasive ventilatory support to avoid tracheotomy indefinitely. Although MIE-EFs can apparently decrease in amyotrophic lateral sclerosis to necessitate tracheotomy, they can increase over time and remain effective in all spinal muscular atrophy types. Two cases demonstrate an association between increasing MIE-EF and, ultimately, successful decannulation of a patient with spinal muscular atrophy type 2 who was continuous tracheostomy mechanical ventilation dependent and a patient with obesity hypoventilation syndrome. Only when MIE-EF increased to exceed 200 l/min did the decannulations succeed. Definitive noninvasive management (continuous noninvasive ventilatory support) of these patients may be possible only when MIE is effective, and the greater the MIE-EF, the greater its effectiveness. Thus, increasing MIE-EF can signal resolution of upper airway obstruction sufficiently to permit decannulation whether a patient is ventilator dependent or not.

Treatment of Idiopathic Diaphragm Flutter: A Case Study.

Diaphragm flutter is a rare disorder defined by dyspnea and often thoracoabdominal pain associated with rapid rhythmic involuntary contractions of the diaphragm with no effective treatment. A 35-year-old woman's flutter was triggered by increasing the depth of breathing and by (electrical) stimulation of the diaphragm. Medical therapy, phrenic nerve crush, and diaphragm pacer stimulation were ineffective. Since increasing diaphragm activity was a trigger, resting the diaphragm was tried. A manual resuscitator and, subsequently, mouthpiece and nasal noninvasive ventilatory support (NVS) instantaneously halted the flutter for 3 months and almost instantaneously for another 6 months. For 16 months, it has continued to halt flutter with rare episodes when getting out of bed that resolve with up to 40 minutes of NVS. To our knowledge, this is the first case of idiopathic diaphragmatic flutter for which diaphragm rest was used as successful treatment with no adverse effects. This should be tried for future cases.

Active lung volume recruitment to preserve vital capacity in Duchenne muscular dystrophy.

OBJECTIVE: To consider the effect of active lung volume recruitment ("air stacking") on rate of decline in vital capacity. DESIGN: Retrospective cross-sectional design. PATIENTS: People with Duchenne muscular dystrophy. METHODS: Vital capacity was measured at every patient visit and then graphed. Air stacking using volume-preset ventilation or manual resuscitator bag was introduced to all patients after their vital capacity plateaued (reached a lifetime maximum). RESULTS: For 151 consecutive patients with multi-year data, the 1-year rate of greatest decline in post-plateau vital capacity was 104 ml (8.8%)/year and occurred from age 20 to 21 years. For 53 patients with multi-visit data for whom air stacking was begun at the immediate post-plateau visit, the 1-year rate of greatest decline in vital capacity was 118 ml (8.5%) and occurred from age 20 to 21 years. Between annual visits, vital capacity and maximum insufflation capacity increased 26.4% and 43.3% of the time, respectively. The peak of maximal vital capacity decline occurred more than 5 years later than previously reported without air stacking. CONCLUSION: For patients with Duchenne muscular dystrophy, active lung volume recruitment may help to preserve vital capacity. Effects on post-plateau vital capacity may be a useful outcome measure for therapeutic trials.

CD40 expression by human melanocytic lesions and melanoma cell lines and direct CD40 targeting with the therapeutic anti-CD40 antibody CP-870,893.

Anti-CD40 antibodies are in clinical development in patients with metastatic melanoma, a cancer that has been reported earlier to express CD40. The antitumor activity of anti-CD40 antibodies may be mediated by direct cytotoxic effects on CD40-positive melanoma cells or indirectly through modulation of host cells. In these studies, biopsies of patients with primary and metastatic melanoma, short-term cultures, and established melanoma cell lines were analyzed for CD40 expression using a combination of methods including immunohistochemistry, flow cytometry, and gene expression profiling, and the cytotoxic effects of the anti-CD40 antibody CP-870,893 on melanoma cell lines were tested using cell viability assays. CD40 was expressed at a higher frequency in metastatic melanoma lesions compared with primary melanomas. There was a variable expression of CD40 in synchronous and metachronous melanoma metastases. Expression of CD40 was present in slightly over half of a large panel of short-term primary melanoma cultures, with a wide range of expression levels by flow cytometry. Similar results were obtained in established melanoma cell lines when analyzed at the mRNA level or by surface protein staining. In approximately one-third of cell lines, the expression of CD40 could be up-regulated with a histone deacetylase inhibitor. Treatment with increasing concentrations of CP-870,893 had no antitumor activity against either CD40-positive or negative melanoma cell lines. In conclusion, approximately one-third to one-half of melanomas expresses CD40 at variable levels. Direct exposure to a CD40-activating antibody does not lead to antitumor activity in melanoma cell lines, suggesting that the antitumor effects of these antibodies in the clinic may be indirectly mediated.

Connective tissue growth factor in regulation of RhoA mediated cytoskeletal tension associated osteogenesis of mouse adipose-derived stromal cells.

BACKGROUND: Cytoskeletal tension is an intracellular mechanism through which cells convert a mechanical signal into a biochemical response, including production of cytokines and activation of various signaling pathways. METHODS/PRINCIPAL FINDINGS: Adipose-derived stromal cells (ASCs) were allowed to spread into large cells by seeding them at a low-density (1,250 cells/cm(2)), which was observed to induce osteogenesis. Conversely, ASCs seeded at a high-density (25,000 cells/cm(2)) featured small cells that promoted adipogenesis. RhoA and actin filaments were altered by changes in cell size. Blocking actin polymerization by Cytochalasin D influenced cytoskeletal tension and differentiation of ASCs. To understand the potential regulatory mechanisms leading to actin cytoskeletal tension, cDNA microarray was performed on large and small ASCs. Connective tissue growth factor (CTGF) was identified as a major regulator of osteogenesis associated with RhoA mediated cytoskeletal tension. Subsequently, knock-down of CTGF by siRNA in ASCs inhibited this osteogenesis. CONCLUSIONS/SIGNIFICANCE: We conclude that CTGF is important in the regulation of cytoskeletal tension mediated ASC osteogenic differentiation.

In vitro expansion of adipose-derived adult stromal cells in hypoxia enhances early chondrogenesis.

Cartilage is an avascular tissue, and chondrocytes in vivo experience a severely hypoxic environment. Using a defined in vitro model of early chondrogenesis, we attempted to enrich for cells with an enhanced ability for chondrogenic differentiation by pre-exposure of mouse adipose-derived adult stromal cells (ADASs) to a hypoxic (2% oxygen) environment. ADASs were subsequently expanded in 2% or 21% oxygen environments, resulting in 2 groups of cells, and then early chondrogenic differentiation was induced at 21% oxygen tension using a 3-dimensional micromass culture system. ADAS chondrogenesis was assessed using Alcian Blue staining for proteoglycans and quantification of sulfated glycosaminoglycans. Osteogenesis of the 2 cell groups was also studied. Two percent oxygen tension profoundly increased the proliferation of ADASs. ADASs expanded in 2% oxygen tension exhibited enhanced early chondrogenic differentiation and diminished osteogenesis, suggesting that the reduced oxygen environment may favor chondroprogenitors. Gene expression analysis suggested that matrix metalloproteinase synthesis was inhibited in cells expanded in 2% oxygen. Furthermore, re-oxygenation of the 2% oxygen-expanded ADASs before differentiation did not significantly affect early chondrogenesis. Thus, priming ADASs with 2% oxygen may have selected for chondrogenic progenitors with an enhanced ability to survive and differentiate. This study is relevant for the future application of cell-based therapies involving cartilage tissue regeneration.

Analysis of the material properties of early chondrogenic differentiated adipose-derived stromal cells (ASC) using an in vitro three-dimensional micromass culture system.

Cartilage is an avascular tissue with only a limited potential to heal and chondrocytes in vitro have poor proliferative capacity. Recently, adipose-derived stromal cells (ASC) have demonstrated a great potential for application to tissue engineering due to their ability to differentiate into cartilage, bone, and fat. In this study, we have utilized a high density three-dimensional (3D) micromass model system of early chondrogenesis with ASC. The material properties of these micromasses showed a significant increase in dynamic and static elastic modulus during the early chondrogenic differentiation process. These data suggest that the 3D micromass culture system represents an in vitro model of early chondrogenesis with dynamic cell signaling interactions associated with the mechanical properties of chondrocyte differentiation.

Hypoxia inducible factor-1alpha deficiency affects chondrogenesis of adipose-derived adult stromal cells.

Increased cartilage-related disease, poor regeneration of cartilage tissue, and limited treatment options have led to intense research in tissue engineering of cartilage. Adipose-derived adult stromal cells (ADAS) are a promising cell source for skeletal tissue engineering; understanding ADAS cellular signaling and chondrogenesis will advance cell-based therapies in cartilage repair. Chondrocytes are unique-they are continuously challenged by a hypoxic microenvironment. Hypoxia inducible factor-1-alpha (HIF-1alpha), a critical mediator of a cell's response to hypoxia, plays a significant role in chondrocyte survival, growth arrest, and differentiation. By using an established in vitro 3-dimensional micromass system, we investigated the role of HIF-1alpha in chondrogenesis. Targeted deletion of HIF-1alpha in ADAS substantially inhibited the chondrogenic pathway specifically. In marked contrast, deletion of HIF-1alpha did not affect osteogenic differentiation but enhanced adipogenic differentiation. This study demonstrates the critical and specific interplay between HIF-1alpha and chondrogenesis in vitro.

Isolation and characterization of posterofrontal/sagittal suture mesenchymal cells in vitro.

BACKGROUND: Craniosynostosis, the premature fusion of cranial sutures, affects one in 2500 children. In the mouse, the posterofrontal suture is programed to fuse postnatally, but the adjacent sagittal suture remains patent throughout life. To study the cellular process of suture fusion, the authors isolated and studied suture-derived mesenchymal cells. METHODS: Skulls were harvested from 80 mice (2 to 5 days old), and posterofrontal and sagittal sutures were dissected meticulously. Suture mesenchymal tissue was separated from the underlying dura mater and overlying pericranium and cultured in growth media. After the cells migrated from the explant tissues, the morphologies of the two cell populations were studied carefully, and quantitative real-time polymerase chain reaction was performed to evaluate gene expression. RESULTS: Both posterofrontal and sagittal cells exhibited highly heterogeneous morphologies, and the posterofrontal cells migrated faster than the sagittal cells. Accordingly, growth factors such as transforming growth factor-beta1 and fibroblast growth factor (FGF)-2 were expressed significantly more highly in posterofrontal compared with sagittal suture mesenchymal cells. In contrast, FGF receptor 2 and FGF-18 were expressed significantly more in sagittal than in posterofrontal suture cells. Importantly, bone morphogenic protein-3, the only osteogenic inhibitor in the bone morphogenic protein family, and noggin, a bone morphogenic protein antagonist, were expressed significantly more in sagittal than in posterofrontal suture cells, suggesting a possible mechanism of suture patency. CONCLUSIONS: To the authors' knowledge, this is the first analysis of mouse suture-derived mesenchymal cells. The authors conclude that isolation of suture-derived mesenchymal cells will provide a useful in vitro system with which to study the mechanisms underlying suture biology.

Mitogenic and chondrogenic effects of fibroblast growth factor-2 in adipose-derived mesenchymal cells.

Adipose-derived mesenchymal cells (AMCs) have demonstrated a great capacity for differentiating into bone, cartilage, and fat. Studies using bone marrow-derived mesenchymal cells (BMSCs) have shown that fibroblast growth factor (FGF)-2, a potent mitogenic factor, plays an important role in tissue engineering due to its effects in proliferation and differentiation for mesenchymal cells. The aim of this study was to investigate the function of FGF-2 in AMC chondrogenic differentiation and its possible contributions to cell-based therapeutics in skeletal tissue regeneration. Data demonstrated that FGF-2 significantly promoted the proliferation of AMCs and enhanced chondrogenesis in three-dimensional micromass culture. Moreover, priming AMCs with treatment of FGF-2 at 10 ng/ml demonstrated that cells underwent chondrogenic phenotypic differentiation, possibly by inducing N-Cadherin, FGF-receptor 2, and transcription factor Sox9. Our results indicated that FGF-2 potentiates chondrogenesis in AMCs, similar to its functions in BMSCs, suggesting the versatile potential applications of FGF-2 in skeletal regeneration and cartilage repair.