Hematopoiesis in mice is extremely resilient to wide variation in TIMP/MMP balance

Tissue inhibitors of matrix metalloproteinases (TIMPs) are natural inhibitors of matrix metalloproteinases (MMPs) and are associated with normal and pathologic extracellular matrix turnover. Because the microenvironment is critical for normal hematopoietic stem/progenitor cell function, we aimed to determine whether alterations in the TIMP/MMP balance impact upon normal hematopoiesis in mice. We have used both overexpression and knockout mouse models to determine whether early hematopoiesis is susceptible to potentially pathologic changes in TIMP/MMP level. These studies used TIMP-1(-/-) mice and retroviral vectors co-expressing human TIMP-1 or TIMP-2 linked with the green fluorescent protein (GFP) transduced into bone marrow (BM) cells and transplanted into lethally-irradiated recipient mice. Loss of TIMP-1 in knockout mice or retroviral overexpression of TIMP-1 or TIMP-2 did not alter hematopoietic stem/progenitor function during steady-state hematopoiesis. Surprisingly, even when applying hematopoietic stress through mobilization, chemotaxis, or myelosuppression, murine hematopoiesis was not adversely affected by TIMP-1 or TIMP-2 level. We conclude that TIMP/MMP balance alone does not exert significant influence on blood cell development and homeostasis. An important corollary of these studies is that specific modulation using MMP inhibitors for cancer or immunologic therapy is unlikely to have adverse hematopoietic side effects.     

Kistrin, a polypeptide platelet GPIIb/IIIa receptor antagonist, enhances and sustains coronary arterial thrombolysis with recombinant tissue-type plasminogen activator in a canine preparation

BACKGROUND. Kistrin is a 68-amino acid polypeptide from the venom of the Malayan pit viper Agkistrodon rhodostoma, which inhibits the platelet GPIIb/IIIa receptor. Its effect on thrombolysis, reocclusion, and bleeding associated with administration of recombinant tissue-type plasminogen activator (rt-PA) was studied in a canine model of coronary artery thrombosis. METHODS AND RESULTS. Coronary patency was monitored for 2 hours by ultrasonic flow probe and repeated coronary angiography. The rt-PA was given as 0.45-mg/kg bolus injections at 15-minute intervals until recanalization or to a maximum of four boluses. Four groups of four or five dogs were studied: a control group that received intravenous heparin (4,000-unit bolus and 1,000 units each hour) and three groups that received heparin and 0.48, 0.24, or 0.12 mg/kg kistrin, administered as a 10% bolus injection and an infusion during a 60-minute period. In the control group, reflow occurred in four of five dogs within 37 +/- 47 minutes but was followed by cyclic reflow and reocclusion. Kistrin at a dose of 0.48 and 0.24 mg/kg reduced the time to reflow to 6 +/- 5 and 10 +/- 3 minutes, respectively, and abolished reocclusion. With 0.12 mg/kg kistrin, reflow occurred in all four animals, within 27 +/- 23 minutes, and reocclusion occurred in two animals. Kistrin induced a dose-related prolongation of the template bleeding time: with 0.48 mg/kg kistrin, the bleeding time was prolonged from 3.8 +/- 1.3 minutes before infusion to 29 +/- 2 minutes during infusion, but it was shortened to 8.3 +/- 2.6 minutes at 90 minutes after the end of infusion. Kistrin also caused a dose-related inhibition of platelet aggregation with ADP and collagen: with 0.48 mg/kg kistrin, platelet aggregation was abolished during the infusion but had partially recovered toward the end of the observation period. Pathological examination of recanalized coronary arterial segments of dogs given 0.48 or 0.24 mg/kg kistrin revealed widely patent arteries with some platelets layered on the damaged intimal surface. CONCLUSIONS. Kistrin increases the rate and extent of thrombolysis with a reduced dose of rt-PA, and it prevents reocclusion. At an effective dose, it is associated with a transient prolongation of the bleeding time and inhibition of platelet aggregation. Kistrin may offer promise as adjunctive treatment to thrombolytic agents in patients with acute myocardial infarction.     

Disposable platform provides visual and color-based point-of-care anemia self-testing

BACKGROUND. Anemia, or low blood hemoglobin (Hgb) levels, afflicts 2 billion people worldwide. Currently, Hgb levels are typically measured from blood samples using hematology analyzers, which are housed in hospitals, clinics, or commercial laboratories and require skilled technicians to operate. A reliable, inexpensive point-of-care (POC) Hgb test would enable cost-effective anemia screening and chronically anemic patients to self-monitor their disease. We present a rapid, stand-alone, and disposable POC anemia test that, via a single drop of blood, outputs color-based visual results that correlate with Hgb levels.METHODS. We tested blood from 238 pediatric and adult patients with anemia of varying degrees and etiologies and compared hematology analyzer Hgb levels with POC Hgb levels, which were estimated via visual interpretation using a color scale and an optional smartphone app for automated analysis.RESULTS. POC Hgb levels correlated with hematology analyzer Hgb levels (r = 0.864 and r = 0.856 for visual interpretation and smartphone app, respectively), and both POC test methods yielded comparable sensitivity and specificity for detecting any anemia (n = 178) (<11 g/dl) (sensitivity: 90.2% and 91.1%, specificity: 83.7% and 79.2%, respectively) and severe anemia (n = 10) (<7 g/dl) (sensitivity: 90.0% and 100%, specificity: 94.6% and 93.9%, respectively).CONCLUSIONS. These results demonstrate the feasibility of this POC color-based diagnostic test for self-screening/self-monitoring of anemia.TRIAL REGISTRATION. Not applicable.FUNDING. This work was funded by the FDA-funded Atlantic Pediatric Device Consortium, the Georgia Research Alliance, Children’s Healthcare of Atlanta, the Georgia Center of Innovation for Manufacturing, and the InVenture Prize and Ideas to Serve competitions at the Georgia Institute of Technology.     

Ultra-protective tidal volume: how low should we go?

Applying tidal volumes of less than 6 mL/kg might improve lung protection in patients with acute respiratory distress syndrome. In a recent article, Retamal and colleagues showed that such a reduction is feasible with conventional mechanical ventilation and leads to less tidal recruitment and overdistension without causing carbon dioxide retention or auto-positive end-expiratory pressure. However, whether the compensatory increase in the respiratory rate blunts the lung protection remains unestablished.Further reducing tidal volumes beyond the standard 6 mL/kg is an appealing goal in patients with acute respiratory distress syndrome (ARDS) [1]. Such reduction could decrease the tidal stretch imposed on the lung, potentially attenuating further the ventilator-induced lung injury [2]. In fact, tidal volumes of less than 6.5 mL/kg and as low as 4 mL/kg were recently associated with increased survival in patients with ARDS [3]. One of the main obstacles to such a strategy is the potential for carbon dioxide (CO₂) retention and severe acidosis. To avoid this, specialized techniques, such as high-frequency oscillatory ventilation and extracorporeal CO₂ removal, have been previously tested with mixed results [4-6].In the previous issue of Critical Care, Retamal and colleagues proposed that lower tidal volumes could be used with conventional positive-pressure ventilation without leading to CO₂ retention [1]. A reduction in tidal volume from 6 to 4 mL/kg was feasible with a decrease in the instrumental dead space and an increase in the respiratory rate. In patients with ARDS, the dead space is a marker of disease severity [7]. Consequently, very low tidal volumes can be difficult to use in practice, especially in very sick patients, because the necessary increase in respiratory rate might cause significant auto-positive end-expiratory pressure (auto-PEEP). Luckily, patients with severe ARDS also tend to have low lung compliance [8], making their lungs inflate and deflate fast. Therefore, this restrictive ventilatory pattern allows the safe use of high respiratory rates without leading to significant auto-PEEP.Retamal and colleagues [1] should be congratulated for their careful design of the ventilator protocol in the 4 mL/kg phase, which allowed an effective CO₂ elimination. The bottom line is that if one decides to use very low tidal volumes with high respiratory rates, attention to the details is invaluable. First, the removal of any dispensable dead space, including substituting an external heated humidifier by the heat-moisture exchanger, is imperative. Second, the use of volume-controlled ventilation helps to keep short inspiratory times. Peak airway pressures may increase, but the preserved expiratory time guarantees low auto-PEEP and, consequently, low plateau pressures. For safety, plateau pressures and auto-PEEP should be measured periodically. Third, in selected cases with high recruitability, the alveolar dead space can be minimized through recruitment maneuvers and higher PEEP values. Finally, the use of a short end-inspiratory pause is encouraged to improve the CO₂ elimination [9]. These measures will improve the safety and optimize the CO₂ elimination of a strategy with very low tidal volumes, even with higher-than-normal respiratory rates.However, even successfully avoiding CO₂ retention, this strategy has yet to be proven effective in terms of further lung protection. We believe that two aspects should be taken into consideration. The first is whether the strategy attenuated the mechanisms of lung injury. The authors performed computed tomography scans in all patients at tidal volumes of both 4 and 6 mL/kg and showed that the amount of cyclic recruitment-derecruitment and hyperinflation decreased after reducing the tidal volume. Although the absolute reduction was small (less than 1% of the lung weight), this finding is suggestive of decreased injury per breath. The second aspect is that an increased respiratory rate can be injurious per se [10]. It would be important to know whether the compensatory increase of the respiratory rate blunted the protective effect per breath of the tidal volume reduction.This tradeoff was emphasized recently in a model of the energy delivered by the ventilator as a surrogate for the potential lung damage [11]. Decreases in tidal volume require disproportionate increases in respiratory rate to maintain alveolar ventilation, and so more energy can be delivered to the lungs even at reduced stress and strain per breath. Though purely theoretical, this hypothesis helps reconcile our expectation of a further protective effect of very low tidal volumes with the recent findings of harmful or null effect of oscillatory high-frequency ventilation [5,6]. In these trials, it is possible that the reduction in lung injury per breath was offset by the very high respiratory rates applied.Finally, Retamal and colleagues [1] followed their patients for 5 to 30 minutes only. Since lower tidal volumes tend to promote atelectasis, especially under insufficient PEEP [12], a longer observation time perhaps would have shown an increase in atelectasis and driving pressures, opposing the benefits initially achieved.In conclusion, we are convinced that a strategy with very low tidal volumes (4 mL/kg) is feasible with conventional positive-pressure ventilation. This strategy could be used in patients with high plateau pressures or high driving pressures with standard 6 mL/kg tidal volumes, but we need more data in terms of lung protection before we can recommend this strategy to every patient with ARDS.     

Rheumatoid arthritis: The diagnostic significance of focal cellular accumulations in the skeletal muscles

Muscle biopsies were obtained in sixteen cases of rheumatoid arthritis in an effort to repeat the work of Freund³ and his collaborators. As controls, biopsies were obtained in fifteen other patients among whom were included cases of osteoarthritis, rheumatic fever, gonococcal arthritis, and “non-specific infectious” arthritis; similar studies were made in twenty-three routine autopsy cases.Muscle lesions were found in thirteen of the sixteen cases of rheumatoid arthritis and in two of the four control cases of “non-specific infectious” arthritis. With the exception of one patient with osteoarthritis who had a history suggestive of rheumatoid arthritis these lesions were absent from all other controls. Our results would give no indication of any fundamental difference between the “atrophic” and the “non-specific infectious” forms of rheumatoid arthritis.The lesions consisted of focal accumulations of lymphocytes and macrophages and occasional plasma cells and eosinophils occurring either in perivascular locations in the perimysium or in the endomysium between the individual muscle fibers. Hyalinization, vacuolization, loss of striations and atrophy of muscle fibers were found frequently in association with these cellular foci in the above group, while in the controls similar degenerative changes occurred without any cellular reaction.These cellular accumulations would appear to be further evidence of the systemic nature of rheumatoid arthritis. Although they have not been found in all cases of this disease, their presence has been limited almost exclusively to rheumatoid arthritis. Their nature is unknown, and their direct relationship to the duration or activity of the disease is as yet undetermined.     

FATE OF THE LYMPHOCYTE

Although the count of circulating lymphocytes in the blood stream remains constant, more lymphocytes enter the blood from the thoracic duct during 24 hours than are present in the blood at any one time. This excess of lymphocytes is not destroyed in the blood stream. The cells migrate from the blood vessels into the mucous membranes and through them to their surface. This occurs chiefly in the gastrointestinal tract, and it is apparently in the mucosa and especially within the intestinal lumen that the function of the lymphocyte is normally performed.     

THE EFFECT OF MECHANICAL TENSION UPON THE POLARITY OF GROWING FIBROBLASTS

These experiments seem to justify the conclusion that mechanical tension may determine the polarity of cell division and line of growth of developing fibroblasts, and further that a shift in tension may cause a shift in position of fibroblasts already formed.