Clinical practice guideline: red blood cell transfusion in adult trauma and critical care.

OBJECTIVE: To develop a clinical practice guideline for red blood cell transfusion in adult trauma and critical care. DESIGN: Meetings, teleconferences and electronic-based communication to achieve grading of the published evidence, discussion and consensus among the entire committee members. METHODS: This practice management guideline was developed by a joint taskforce of EAST (Eastern Association for Surgery of Trauma) and the American College of Critical Care Medicine (ACCM) of the Society of Critical Care Medicine (SCCM). We performed a comprehensive literature review of the topic and graded the evidence using scientific assessment methods employed by the Canadian and U.S. Preventive Task Force (Grading of Evidence, Class I, II, III; Grading of Recommendations, Level I, II, III). A list of guideline recommendations was compiled by the members of the guidelines committees for the two societies. Following an extensive review process by external reviewers, the final guideline manuscript was reviewed and approved by the EAST Board of Directors, the Board of Regents of the ACCM and the Council of SCCM. RESULTS: Key recommendations are listed by category, including (A) Indications for RBC transfusion in the general critically ill patient; (B) RBC transfusion in sepsis; (C) RBC transfusion in patients at risk for or with acute lung injury and acute respiratory distress syndrome; (D) RBC transfusion in patients with neurologic injury and diseases; (E) RBC transfusion risks; (F) Alternatives to RBC transfusion; and (G) Strategies to reduce RBC transfusion. CONCLUSIONS: Evidence-based recommendations regarding the use of RBC transfusion in adult trauma and critical care will provide important information to critical care practitioners.

Diethylene glycol monomethyl ether, ethylene glycol monomethyl ether and the metabolite, 2-methoxyacetic acid affect in vitro chondrogenesis.

Diethylene glycol monomethyl ether (DEGME), ethylene glycol monomethyl ether (EGME) and their common metabolite, methoxyacetic acid (MAA) have been associated with adverse reproductive effects. The objective of this research is to investigate the effects of DEGME, EGME and MAA on in vitro chondrogenesis and the mechanisms by which these effects occur. Micromass cultures were exposed to DEGME, EGME or MAA for 5 days and proteoglycan abundance and cell proliferation determined. Longer-term 9- and 14-day cultures were exposed to MAA and apoptosis analyzed. All three chemicals decreased proteoglycan abundance and cell proliferation at the highest dose tested (100 microL/mL). However, only MAA showed a dose-dependent effect for both parameters at 0.01, 10, and 100 microL/mL. Furthermore, micromass cultures show an increase in apoptotic cells which when treated with MAA suggest that cell death could result from induced apoptosis. These results suggest that effects of DEGME and EGME are the result of generalized toxicity, but their metabolite MAA induces mitochondrial-mediated apoptosis during in vitro chondrogenesis.

Lymphadenectomy prior to rat hind limb allotransplantation prevents graft-versus-host disease in chimeric hosts.

In previous rat studies, the use of mixed allogeneic chimerism (MAC) to induce host tolerance to hind limb allografts has resulted in severe graft-versus-host disease (GVHD). The purpose of this study was to determine if immunocompetent cells in bone marrow (BM) and/or lymph nodes (LNs) of transplanted limbs were responsible for inducing GVHD in mixed chimeric hosts. [ACI-->Wistar Furth] chimeric rats received ACI hind limbs that were non-irradiated, irradiated (1050 cGy) or lymphadenectomized. Rejection, GVHD and donor chimerism was assessed. Chimeric hosts rejected none of their limbs. However, hosts of non-irradiated hind limbs succumbed to GVHD 22.4+/-0.8 days after transplantation. In contrast, chimeras that received irradiated or lymphadenectomized ACI hind limbs showed no clinical or histological signs of GVHD at 5 months. We conclude that mixed chimeric hosts are susceptible to GVHD due to the immunocompetent cell load provided by the LNs, not the BM, of hind limb allografts.

Composite tissue allotransplantation in chimeric hosts: part I. Prevention of graft-versus-host disease.

BACKGROUND: Mixed allogeneic chimerism (MAC) has been shown to induce tolerance to composite tissue allografts (CTA). However, transplantation of unmanipulated donor-specific limbs results in severe graft-versus-host disease (GVHD). This suggests that nontolerant mature donor-derived cells in the CTA may affect the stability of chimerism, potentially resulting in GVHD. The aim of this study was to develop an approach to study and prevent GVHD in a mixed chimeric-rat hind-limb transplantation model. METHODS: [ACI-->WF] chimeras received a limb from Wistar Furth (WF) (syngeneic), Fisher (third-party), or ACI (irradiated [1,050 cGy] or nonirradiated) rats. In vitro tolerance was assessed using mixed lymphocyte reactivity (MLR) assays at the time the animals were killed. RESULTS: [ACI-->WF] chimeras with greater than 85% chimerism exhibited rejection-free survival of donor-specific hind limbs. However, 100% of these animals developed lethal GVHD 22.4+/-2.8 days after limb transplantation. [ACI-->WF] chimeras that underwent transplantation with irradiated ACI or syngeneic WF limbs showed no signs of rejection or GVHD at 5 months. Nonchimeric and third-party controls rejected limbs within 10 days. CONCLUSIONS: Conditioning of the host WF rats with 950 cGy of irradiation (sublethal, myeloablative) led to high levels of MAC without GVHD. The mature T-cell content of nonirradiated donor (ACI) limbs was sufficient to induce lethal GVHD in 100% of tolerant mixed chimeric [ACI-->WF] hosts. Irradiation of donor limbs before transplantation resulted in long-term donor-specific tolerance and prevented GVHD. These data demonstrate that (1) established chimeras could be susceptible to GVHD caused by immunocompetent donor cells transferred with the hind limb, and (2) inactivating these cells with irradiation prevents GVHD and destabilization of chimerism, and permits rejection-free graft acceptance.

Composite tissue allotransplantation in chimeric hosts part II. A clinically relevant protocol to induce tolerance in a rat model.

BACKGROUND: We and others have shown that mixed allogeneic chimerism induces donor-specific tolerance to composite tissue allografts across major histocompatibility complex barriers without the need for immunosuppression. However, a delay period between bone marrow transplantation and limb allotransplantation is required, making such protocols impractical for clinical application. This study eliminates this delay period in a rat hind limb allotransplantation model by performing mixed allogeneic chimerism induction and transplantation "simultaneously." METHODS: Group 1 included controls in which naïve Wistar Furth (WF) hosts received ACI hind limbs. Group 2 included (ACI-->WF) chimeras that received limbs from third-party donors (Fisher), and group 3 included chimeras that received irradiated (1,050 cGy) ACI limbs. In group 4, WF hosts conditioned with 950 cGy received irradiated (1,050 cGy) ACI limbs followed by infusion of 100 x 10(6) ACI T-cell-depleted bone marrow cells and immunotherapy (tacrolimus and mycophenolate mofetil) for 28 days. Group 5 animals received the same treatment as group 4 animals without immunotherapy. RESULTS: The rats in groups 1 and 2 rejected their limbs within 10 days. Only one rat in group 4 survived to the end of the study. Groups 3 and 5 demonstrated long-term limb survival without rejection or graft-versus-host disease. High levels of donor chimerism (>80%) were achieved and maintained throughout the study. Mixed lymphocyte reaction assays in both groups revealed donor-specific hyporesponsiveness with vigorous third-party reactivity. CONCLUSIONS: This study demonstrated that infusion of donor bone marrow cells into conditioned hosts immediately after limb transplantation results in stable mixed chimerism, robust tolerance, and reliable limb allograft survival.

Can angiogenesis induced by chronic electrical stimulation enhance latissimus dorsi muscle flap survival for application in cardiomyoplasty?

In cardiomyoplasty, the latissimus dorsi muscle is lifted on its primary neurovascular pedicle and wrapped around a failing heart. After 2 weeks, it is trained for 6 weeks using chronic electrical stimulation, which transforms the latissimus dorsi muscle into a fatigue-resistant muscle that can contract in synchrony with the beating heart without tiring. In over 600 cardiomyoplasty procedures performed clinically to date, the outcomes have varied. Given the data obtained in animal experiments, the authors believe these variable outcomes are attributable to distal latissimus dorsi muscle flap necrosis. The aim of the present study was to investigate whether the chronic electrical stimulation training used to transform the latissimus dorsi muscle into fatigue-resistant muscle could also be used to induce angiogenesis, increase perfusion, and thus protect the latissimus dorsi muscle flap from distal necrosis. After 14 days of chronic electrical stimulation (10 Hz, 330 microsec, 4 to 6 V continuous, 8 hours/day) of the right or left latissimus dorsi muscle (randomly selected) in 11 rats, both latissimus dorsi muscles were lifted on their thoracodorsal pedicles and returned to their anatomical beds. Four days later, the resulting amount of distal flap necrosis was measured. Also, at predetermined time intervals throughout the experiment, muscle surface blood perfusion was measured using scanning laser Doppler flowmetry. Finally, latissimus dorsi muscles were excised in four additional stimulated rats, to measure angiogenesis (capillary-to-fiber ratio), fiber type (oxidative or glycolytic), and fiber size using histologic specimens. The authors found that chronic electrical stimulation (1) significantly (p < 0.05) increased angiogenesis (mean capillary-to-fiber ratio) by 82 percent and blood perfusion by 36 percent; (2) did not reduce the amount of distal flap necrosis compared with nonchronic electrical stimulation controls (29 +/- 5.3 percent versus 26.6 +/- 5.1 percent); (3) completely transformed the normally mixed (oxidative and glycolytic) fiber type distribution into all oxidative fibers; and (4) reduced fiber size in the proximal and middle but not in the distal segments of the flap. Despite the significant increase in angiogenesis and blood perfusion, distal latissimus dorsi muscle flap necrosis did not decrease. This might be because of three reasons: first, the change in muscle metabolism from anaerobic to aerobic may have rendered the muscle fibers more susceptible to ischemia. Second, because of the larger diameter of the distal fibers in normal and stimulated latissimus dorsi muscle, the diffusion distance for oxygen to the center of the distal fibers is increased, making fiber survival more difficult. Third, even though angiogenesis was significantly increased in the flap, cutting all but the single vascular pedicle resulted in the newly formed capillaries not receiving enough blood to provide nourishment to the distal latissimus dorsi muscle. The authors' findings indicate that chronic electrical stimulation as tested in these experiments could not be used to prevent distal latissimus dorsi muscle flap ischemia and necrosis in cardiomyoplasty.

The effects of a broad-spectrum matrix metalloproteinase inhibitor on characteristics of wound healing.

This study investigates the effects of a broad-spectrum matrix metalloproteinase inhibitor (MMP-i) on the rate of closure, hydroxyproline deposition, and macrophage infiltration in healing wounds. Full-thickness excisional wounds were created on the dorsal surface of hairless mice. Two experimental groups were used to measure rates of wound closure: (a) MMP-i administration (0.03, 0.3, 3.0, and 30 microg/mL) on days 0-1 postwounding (inflammatory phase) and (b) MMP-i administration (0.03, 0.3, 3.0, and 30 microg/mL) on days 6-8 postwounding (proliferative phase). Additionally, hydroxyproline deposition and percent macrophage infiltration were measured in skin wound margins on days 2, 8, and 16 postwounding. MMP-i administration at concentrations of 0.03, 0.3, and 3.0 microg/mL on days 0-1 postwounding significantly (p <.05) increased the rate of wound closure. No significant effect on the rate of wound closure was observed with MMP-i administration on days 6-8 postwounding. Hydroxyproline deposition was significantly (p <.05) increased on day 8 postwounding, and the percent macrophage infiltration was significantly (p <.05) decreased on day 2 postwounding by MMP-i administration on days 0-1 postwounding. These experiments demonstrate that MMP-i administration during the inflammatory phase significantly affects several characteristics of wound healing. We postulate that these effects may be attributed to decreased degradation of ECM components, increased concentrations of endogenous growth factors, and a shortened inflammatory phase.

Role of PKC in the late phase of microvascular protection induced by preconditioning.

INTRODUCTION: We hypothesized that the late phase of microvascular protection induced by ischemic preconditioning or by adenosine is protein kinase C (PKC) dependent. MATERIALS AND METHODS: The cremaster muscle of male Sprague-Dawley rats underwent 45 min of ischemic preconditioning and, 24 h later, 4 h of warm ischemia followed by 60 min of reperfusion. To mimic the effects of IPC, adenosine (ADO; an adenosine receptor agonist) or 4-phorbol 12-myristate 13-acetate (PMA; a PKC activator) was delivered to the vascular network of the cremaster 24 h before the prolonged ischemia via local intra-arterial infusion. To block the microvascular protection induced by ADO or IPC, chelerythrine (CHE; a PKC blocker) was given by local intra-arterial infusion prior to the administration of ADO or the initiation of IPC. Microvascular responses in the cremaster muscle to ischemic preconditioning or pharmacological preconditioning were determined by measuring terminal arteriole diameter and capillary perfusion using intravital microscopy and by the evaluation of the endothelium-dependent nitric oxide system in terminal arterioles. RESULTS: Blockade of PKC using CHE on day 1 eliminated both ADO- and IPC-induced microvascular protections seen on day 2. However, the microvascular protection induced by the administration of PMA (without IPC) that was given 24 h before the 4 h of warm ischemia/reperfusion was significantly better than the control group response (sham IPC), but was not as good as the protection induced by IPC or ADO alone. CONCLUSION: The overall results from these studies suggest that ischemic or ADO preconditioning induces late-phase microvascular protection in skeletal muscle by a PKC-dependent mechanism.

Reducing the vascular delay period in latissimus dorsi muscle flaps for use in cardiomyoplasty.

Although the mechanism by which vascular delay benefits skin flaps is not completely understood, this topic has been extensively studied and reported on in the literature. In contrast, little has been documented about the effects of vascular delay in skeletal muscle flaps. Recent animal studies tested the effectiveness of vascular delay to enhance latissimus dorsi muscle flap viability for use in cardiomyoplasty and found that it prevented distal flap necrosis. However, these studies did not define the optimal time period necessary to achieve this beneficial effect. The purpose of this study was to determine how many days of "delay" can elicit the beneficial effects of vascular delay on latissimus dorsi muscle flaps. To accomplish this, 90 latissimus dorsi muscles of 45 male Sprague-Dawley rats were randomly subjected to vascular delay on one side or a sham procedure on the other. After predetermined delay periods (0, 3, 7, 10, and 14 days) or a sham procedure, all latissimus dorsi muscles were elevated as single pedicled flaps based only on their thoracodorsal neurovascular pedicle. Latissimus dorsi muscle perfusion was measured using a Laser Doppler Perfusion Imager just before and immediately after flap elevation. The muscles were then returned to their original vascular beds, isolated from adjacent tissue with Silastic film, sutured into place to maintain their original size and shape, and left there for 5 days. After 5 days, the latissimus dorsi muscle flaps were dissected free, scanned again (Laser Doppler Perfusion Imager-perfusion measurements), and the area of distal necrosis was measured using digitized planimetry of magnified images. The authors' results showed that delay periods of 3, 7, 10, and 14 days significantly increased (p < 0.05) blood perfusion and decreased (p < 0.05) distal flap necrosis when compared with sham controls. On the basis of these findings, the authors conclude that in their rat latissimus dorsi muscle flap model the beneficial effects of vascular delay are present as early as 3 days. If these findings also hold true in humans, they could be useful in cardiomyoplasty by allowing surgeons to shorten the amount of time between the vascular delay procedure and the cardiomyoplasty procedure in these very sick patients.