Cold Preservation Research Articles

P 105 - A lesson from the oxidative metabolism of a hibernator's heart: strategy for cardioprotection

Maintaining heart viability in states of unfavorable metabolite/oxygen supply has become a prime challenge in cardiac medicine. Mitochondria have been recognised as key actors in cardioprotection. Our goal was to reveal the myocardial protective phenotype related to mitochondrial bioenergetic pathways in the state of limited metabolite supply and low temperature using the physiological model of mammalian hibernation. European ground squirrels (Spermophilus citellus) were exposed to cold (4 ± 1 °C) and divided into two groups: (1) animals that fell into torpor (hibernating group) and (2) animals that stayed active and euthermic for 1, 3, 7, 12, or 21 days (cold-exposed group). We found increased protein levels of electron transport chain components and ATP synthase in late cold acclimation and in hibernation. Also, protein level of PGC-1α was upregulated. Phospho-AMPKα protein content was increased during early cold acclimation. HIF1α protein level was unchanged. Protein levels of manganese and copper-zinc superoxide dismutase and glutathione peroxidase were increased in hibernation. The hibernation related phenotype of the heart is characterised by controlled improvement of mitochondrial energy and antioxidative capacity. Its reproduction could have broad implications, from myocardial protection in ischaemia/reperfusion to the hypothermic survival and cold preservation of organs.

Role of temperature in reconditioning and evaluation of cold preserved kidney and liver grafts.

Purpose of reviewOrgan shortage in transplantation medicine forces surgical research toward the development of more efficient approaches in organ preservation to enable the application of ‘less than optimal’ grafts. This review summarizes current techniques aiming to recondition cold-stored organ grafts prior to transplantation to reduce reperfusion-induced tissue injury and improve postimplantation graft function.Recent findingsEnd-ischemic reconditioning has classically been attempted by cold oxygenated perfusion. By contrast, evaluation of graft performance prior to transplantation might be facilitated by perfusion at higher temperatures, ideally at normothermia. A drastic temperature shift from cold preservation to warm perfusion, however, has been incriminated to trigger a so-called rewarming injury associated with mitochondrial alterations. A controlled gradual warming up during machine perfusion could enhance the restitution of cellular homeostasis and improve functional outcome upon warm reperfusion.SummaryMachine perfusion after conventional cold storage is beneficial for ulterior function after transplantation. Cold grafts should be initially perfused at low temperatures allowing for restitution of cellular homeostasis under protective hypothermic limitation of metabolic turnover. Delayed slow rewarming of the organ might further mitigate rewarming injury upon reperfusion and also increases the predictive power of evaluative measures, taken during pretransplant perfusion.

A safety evaluation of profound hypothermia-induced suspended animation for delayed resuscitation at 90 or 120\xa0min

BackgroundThe successful treatment of military combat casualties with penetrating injuries is significantly dependent on the time needed to get the patient to an adequate treatment facility. Profound hypothermia-induced suspended animation for delayed resuscitation (SADR) is a novel approach for inducing cardiac arrest and buying additional time for such injuries. However, the time used to safely administer circulatory arrest (CA) is controversial. The goal of this study was to evaluate the safety of hypothermia-induced SADR over 90 and 120 min time intervals.MethodsSixteen male BAMA minipigs were randomized into two groups: CA90 group (90 min, n = 8) and CA120 group (120 min, n = 8). Cannulation of the right common carotid arteries and internal jugular veins was performed to establish cardiopulmonary bypass for each animal. Through the perfusion of cold organ preservation solution (OPS), cardioplegia and profound hypothermia (15 °C) were induced. After CA, cardiopumonary bypass (CPB) was restarted, and the animals were gradually re-warmed and resuscitated. The animals were assisted with ventilators until spontaneous breathing was achieved. The index of hemodynamic perioperative serum chemistry values [alanine transaminase (ALT), aspartate aminotransferase (AST), creatinine (CR), lactic dehydrogenase (LDH) and troponin T (TnT)] and survival were observed from pre-operation to 7 days post-operation.ResultsFifteen animals were enrolled in the experiment, while 1 animal in CA120 group died from surgical error. All 8 animals in CA90 group recovered, with only 1 animal displaying mild disability. However, in CA120 group, only 2 animals survived with severe disability, and the other 5 animals died after 2 days post-operation. In CA90 group, the perioperative serum chemistry values increased at 1 day post-operation (ALT 84.43 ± 18.65 U/L; AST 88.99 ± 23.19 U/L; Cr 87.90 ± 24.49 μmol/L; LDH 1894.13 ± 322.26 U/L; TnT 0.849 ± 0.135 ng/ml) but decreased to normal or almost normal levels at 7 days post-operation (ALT 52.48 ± 9.04 U/L; AST 75.23 ± 21.46 U/L; Cr 82.69 ± 18.41 μmol/L; LDH 944.67 ± 834.32 U/L; TnT 0.336 ± 0.076 ng/ml).ConclusionsProfound hypothermia-induced SADR is an effective method for inducing cardiac arrest. Our results indicate that inducing CA for 90 min (at 15 °C) is safer than doing so for 120 min. Our results indicate that 120 min of CA at 15 °C is dangerous and can result in high mortality and severe neurological complications. Further experimentation is needed to determine whether 120 min of CA at temperatures lower than 15 °C can lead to safe recovery.

Reduced-cost Chlamydia trachomatis-specific multiplex real-time PCR diagnostic assay evaluated for ocular swabs and use by trachoma research programmes

IntroductionTrachoma, caused by the intracellular bacterium Chlamydia trachomatis (Ct), is the leading infectious cause of preventable blindness. Many commercial platforms are available that provide highly sensitive and specific detection of Ct DNA. However, the majority of these commercial platforms are inaccessible for population-level surveys in resource-limited settings typical to trachoma control programmes. We developed two low-cost quantitative PCR (qPCR) tests for Ct using readily available reagents on standard real-time thermocyclers. MethodsEach multiplex qPCR test targets one genomic and one plasmid Ct target in addition to an endogenous positive control for Homo sapiens DNA. The quantitative performance of the qPCR assays in clinical samples was determined by comparison to a previously evaluated droplet digital PCR (ddPCR) test. The diagnostic performance of the qPCR assays were evaluated against a commercial assay (artus C. trachomatis Plus RG PCR, Qiagen) using molecular diagnostics quality control standards and clinical samples. We examined the yield of Ct DNA prepared from five different DNA extraction kits and a cold chain-free dry-sample preservation method using swabs spiked with fixed concentrations of human and Ct DNA. ResultsThe qPCR assay was highly reproducible (Ct plasmid and genomic targets mean total coefficients of variance 41.5% and 48.3%, respectively). The assay detected 8/8 core specimens upon testing of a quality control panel and performed well in comparison to commercially marketed comparator test (sensitivity and specificity>90%). Optimal extraction and sample preservation methods for research applications were identified. ConclusionWe describe a pipeline from collection to diagnosis providing the most efficient sample preservation and extraction with significant per test cost savings over a commercial qPCR diagnostic assay. The assay and its evaluation should allow control programs wishing to conduct independent research within the context of trachoma control, access to an affordable test with defined performance characteristics.

Lungs donated after circulatory death and prolonged warm ischemia are transplanted successfully after enhanced ex vivo lung perfusion using adenosine A2B receptor antagonism

ObjectiveThe current supply of acceptable donor lungs is not sufficient for the number of patients awaiting transplantation. We hypothesized that ex vivo lung perfusion (EVLP) with targeted drug therapy would allow successful rehabilitation and transplantation of donation after circulatory death lungs exposed to 2 hours of warm ischemia. MethodsDonor porcine lungs were procured after 2 hours of warm ischemia postcardiac arrest and subjected to 4 hours of cold preservation or EVLP. ATL802, an adenosine A2B receptor antagonist, was administered to select groups. Four groups (n = 4/group) were randomized: cold preservation (Cold), cold preservation with ATL802 during reperfusion (Cold + ATL802), EVLP (EVLP), and EVLP with ATL802 during ex vivo perfusion (EVLP + ATL802). Lungs subsequently were transplanted, reperfused, and assessed by measuring dynamic lung compliance and oxygenation capacity. ResultsEVLP + ATL802 significantly improved dynamic lung compliance compared with EVLP (25.0 ± 1.8 vs 17.0 ± 2.4 mL/cmH2O, P = .04), and compared with cold preservation (Cold: 12.2 ± 1.3, P = .004; Cold + ATL802: 10.6 ± 2.0 mL/cmH2O, P = .002). Oxygenation capacity was highest in EVLP (440.4 ± 37.0 vs Cold: 174.0 ± 61.3 mm Hg, P = .037). No differences in oxygenation or pulmonary edema were observed between EVLP and EVLP + ATL802. A significant decrease in interleukin-12 expression in tissue and bronchoalveolar lavage was identified between groups EVLP and EVLP + ATL802, along with less neutrophil infiltration. ConclusionsSeverely injured donation after circulatory death lungs subjected to 2 hours of warm ischemia are transplanted successfully after enhanced EVLP with targeted drug therapy. Increased use of lungs after uncontrolled donor cardiac death and prolonged warm ischemia may be possible and may improve transplant wait list times and mortality.

Role of MitoBK Channels in Renal Cold Preservation

Patients with end‐stage renal failure require kidney transplantation to avoid life‐long hemodialysis associated with high mortality. Kidneys from brain‐dead or deceased donors are placed in cold storage (CS) preservation solutions until a recipient is identified. However, kidneys exposed to CS solutions are subjected to extended ischemia, and our laboratory reported that renal CS induces oxidative stress and subsequent mitochondrial damage. Thus, there is a critical need to develop strategies to protect donor kidneys from CS ‐induced damage to improve long‐term renal function in the transplant recipients. In this regard, the high‐conductance Ca2+‐activated K+ (BK) channel recently was discovered in the mitochondrial inner membrane (mitoBK) of cardiomyocytes and other cell types. When activated, the mitoBK channel reportedly protects against ischemia‐reperfusion injury in the heart by reducing mitochondrial damage and reactive oxygen species (ROS). We show, for the first time, that the mitoBK channel is expressed in the mitochondria of rat renal proximal tubular cells in vitro and in rat kidney in vivo. Our preliminary evidence suggests that CS reduces expression of the pore‐forming a subunit of the mitoBK channel in rat renal proximal tubular cell (n=3) and rat kidney (n=4) whole‐cell lysates. Thus, we hypothesize that: MitoBK channels down‐regulate in response to CS‐induced mitochondrial damage, contributing to donor kidney injury. We also propose that restoring mitoBK channel function will provide protection from CS ‐induced injury and improve transplanted kidney function. Our preliminary studies using High Resolution Respirometry (HRR) with fresh kidney biopsies show that Complex I, II, and III (but not Complex IV) activity was significantly impaired in rat kidneys exposed to CS for 18 hours. Twenty‐four hours after renal transplant, recipients of kidneys exposed to 18 hours of CS showed elevated serum creatinine (4.4 mg/dl, n=6) compared to rats having undergone only sham surgery (0.3 mg/dl, n=5). Blood urea nitrogen also was elevated in transplant recipients of CS kidneys (CS + transplant, ~140 mg/dl; sham, ~20 mg/dl). These indicators of impaired renal function correlated with loss of mitoBK protein in CS kidneys. Ongoing patch‐clamp studies are evaluating the properties and function of mitoBK channels in our cell and animal models of renal CS, and evaluating whether pharmacological activation of mitoBK channels during CS can attenuate ROS, limit mitochondrial injury, and improve the function of transplanted kidneys.Support or Funding InformationFunded by AHA Predoctoral Fellowship #16PRE30830010 and UAMS pilot grant G1‐51454‐99.

Improved metabolism and redox state with a novel preservation solution: implications for donor lungs after cardiac death (DCD).

Lungs donated after cardiac death (DCD) are an underutilized resource for a dwindling donor lung transplant pool. Our study investigates the potential of a novel preservation solution, Somah, to better preserve statically stored DCD lungs, for an extended time period, when compared to low-potassium dextran solution (LPD). We hypothesize that Somah is a metabolically superior organ preservation solution for hypothermic statically stored porcine DCD lungs, possibly improving lung transplant outcomes. Porcine DCD lungs (n = 3 per group) were flushed with and submerged in cold preservation solution. The lungs were stored up to 12 h, and samples were taken from lung tissue and the preservation medium throughout. Metabolomic and redox potential were analyzed using high performance liquid chromatography, mass spectrometry, and RedoxSYS®, comparing substrate and pathway utilization in both preservation solutions. Glutathione reduction was seen in Somah but not in LPD during preservation. Carnitine, carnosine, and n-acetylcarnosine levels were elevated in the Somah medium compared with LPD throughout. Biopsies of Somah exposed lungs demonstrated similar trends after 2 h, up to 12 h. Adenosine gradually decreased in Somah medium over 12 h, but not in LPD. An inversely proportional increase in inosine was found in Somah. Higher oxidative stress levels were measured in LPD. Our study suggests suboptimal metabolic preservation in lungs stored in LPD. LPD had poor antioxidant potential, cytoprotection, and an insufficient redox potential. These findings may have immediate clinical implications for human organs; however, further investigation is needed to evaluate DCD lung preservation in Somah as a viable option for transplant.

Prolonging hypothermic ischaemic cardiac and vascular storage by inhibiting the activation of the nuclear enzyme poly(adenosine diphosphate-ribose) polymerase.

Heart transplantation is the standard treatment in end-stage heart failure and at shortage of cardiac allografts is its major limiting factor. Striving to optimize the use of this limited resource, the aspect that long distance procurement may increase the available donor pool must be taken into consideration. As poly(ADP-ribose)polymerase (PARP)-activation has been identified as a key pathway of reperfusion injury, we assessed the hypothesis that its inhibition would allow an extension of cold preservation time and protect the graft against ischaemia/reperfusion injury. Hearts from donor rats were explanted, stored in a preservation solution (Custodiol) at 4 °C for 4 h or 8 h, and heterotopically transplanted. A vehicle or the PARP-inhibitor, INO-1001 (5 mg/kg), was administered during the reperfusion period. We evaluated post-transplant graft function with a Millar micromanometer at different left-ventricular volumes. Additionally, in organ bath experiments the effect of PARP-inhibition on endothelium-dependent and -independent vasorelaxation was evaluated after long-term cold ischaemic storage/warm reperfusion. PARP-inhibition resulted in a better systolic functional recovery of grafts submitted to 4 h and 8 h ischaemia. Furthermore, INO-1001 decreased the left-ventricular end-diastolic pressure after 8 h of ischaemia. Coronary blood flow was significantly higher after PARP-inhibition in comparison to controls. Endothelium-dependent vasorelaxation was significantly better in the INO-1001-groups than in the vehicle-treated transplant groups. After 24-h hypothermic storage, treatment of aortic ring with INO-1001 during reoxygenation significantly improved endothelial dysfunction. By inhibiting the PARP activation, INO-1001 can protect the graft and endothelium from the injury that is caused by prolonged cold myocardial ischaemia/reperfusion, thereby improving post-transplant graft function.

Effect of Hypothermic Machine Perfusion on the Preservation of Kidneys Donated After Cardiac Death: A Single-Center, Randomized, Controlled Trial.

To assess the application of a hypothermic machine perfusion device (LifePort) in kidney transplantation from donation after cardiac death (DCD) donors, 24 pairs of DCD kidneys were randomly divided into two groups: one of the paired kidneys from the same donor was perfused with the LifePort machine (hypothermic machine perfusion [HMP]), and the contralateral kidney was prepared using common static cold preservation (CCP). The two groups were compared with respect to the incidence of delayed graft function (DGF), level of graft function, and pathological changes in time-zero biopsy specimens. The incidence of DGF was 16.7 and 37.5% in the HMP and CCP groups, respectively; the difference between the two groups was statistically significant (P < 0.05). The incidence of acute rejection was 4.1 (1/24) and 8.3% (2/24) in the HMP and CCP groups, respectively; this difference was not statistically significant (P > 0.05). Forty-eight kidney patients were followed up for 6 months, and the two groups of recipients all survived, yielding a survival rate of 100%. The mean 6-month serum creatinine levels were 98.7 ± 23.6 µmol/L in the HMP group and 105.3 ± 35.1 µmol/L in the CCP group; there was no significant difference between the two groups. HMP can reduce the incidence of DGF in DCD kidneys, and this effect is greater for expanded criteria donors kidneys. HMP can also improve early renal function.

A Soluble Form of P Selectin Glycoprotein Ligand 1 Requires Signaling by Nuclear Factor Erythroid 2-Related Factor 2 to Protect Liver Transplant Endothelial Cells Against Ischemia-Reperfusion Injury.

Liver endothelial cell (LEC) damage is essential in the pathogenesis of ischemia-reperfusion injury (IRI) in transplant recipients. We analyzed the mechanism of LEC resistance against IRI by using a novel recombinant soluble form of P selectin glycoprotein ligand 1, tandem P selectin glycoprotein ligand immunoglobulin (TSGL-Ig), in a mouse model of hepatic cold preservation (4°C in University of Wisconsin solution for 20 h) and syngeneic orthotopic liver transplantation (OLT). Unlike controls, TSGL-Ig protected orthotopic liver transplants against ischemia-reperfusion (IR) stress, shown by depressed serum alanine aminotransferase levels, well-preserved hepatic architecture, and improved survival (42% vs. 92%). TSGL-Ig suppressed neutrophil/macrophage sequestration and proinflammatory cytokine/chemokine programs in OLT. Treatment with TSGL-Ig mitigated LEC activation (P and E selectin, VCAM-1 and intercellular adhesion molecule 1 expression). In parallel in vitro studies, TSGL-Ig diminished cellular damage in H2 O2 -stressed LEC cultures (lactic acid dehydrogenase and alanine aminotransferase levels). Increased thioredoxin, glutamate-cysteine ligase, NAD(P)H quinone dehydrogenase 1, and hypoxia-inducible factor 1α expression, along with transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2), implied that TSGL-Ig exerts antioxidant functions in IR-stressed OLT and H2 O2 -stressed LECs. Indeed, Nrf2-deficient livers suffered fulminant IRI compared with WT despite concomitant TSGL-Ig therapy. Thus, TSGL-Ig is not only acting as a competitive antagonist blocking leukocyte migration into IR-stressed liver, but it may also act directly as an agonist stimulating Nrf2-mediated cytoprotection in LECs. This study supports the role of P selectin signaling in hepatic homeostasis in OLT, with broad implications for tissue damage conditions.

Analysis about Heat Transfer of Vegetables during Cold Shock Treatment and Preservation Quality after Storage

Abstract This study investigated the heat transfer mechanism and preservation quality of axisymmetric vegetables during cold shock treatment (CST). A simulation model of heat transfer was developed based on geometrical and biological characteristics of the vegetables by finite element difference method. CST for cucumbers and carrots with cold water at 0°C (40 min), 2°C (20, 40 and 60 min) and 4°C (40 min) was conducted to evaluate the relationships between heat transfer characteristics and preservation quality. The comparative analysis shown that the simulated results agreed well with measured results. Meanwhile, in terms of weight loss, firmness and POD activity, CST with cold water at 2°C for 40 min could achieve the most positive effect on preserving cucumbers. The cucumber during treatment experienced whole variable temperature stage and appropriate constant temperature stage so as to reach optimal preservation quality.

Activation of Fibrinolysis, But Not Coagulation, During End-Ischemic Ex Situ Normothermic Machine Perfusion of Human Donor Livers.

Ex situ normothermic machine perfusion (NMP) can be performed after traditional static cold preservation to assess graft function and viability before transplantation. It is unknown whether this results in activation of coagulation and fibrinolysis, as may occur upon graft reperfusion in vivo. Twelve donor livers declined for transplantation underwent 6 hours of end-ischemic NMP using a heparinized plasma-based perfusion fluid. Concentration of prothrombin fragment F1 + 2 (marker of coagulation activation), D-dimer, plasmin-antiplasmin complex, tissue plasminogen activator and plasminogen activator inhibitor-1 (markers for fibrinolysis) and alanine aminotransferase (ALT) (marker of ischemia-reperfusion [I/R] injury) were measured in perfusion fluid at regular intervals. Liver biopsies were examined for the presence of fibrin, using light microscopy after Maurits, Scarlet and Blue staining. No significant increase in prothrombin F1 + 2 was noted during NMP. D-dimer and plasmin-antiplasmin complex levels increased soon after start of NMP and D-dimer concentrations correlated significantly with levels of tissue plasminogen activator. In livers displaying good function during NMP, perfusate levels of ALT and D-dimers were low (≤3500 ng/mL), whereas significantly higher D-dimer levels (>3500 ng/mL) were in found in livers with poor graft function. Activation of fibrinolysis correlated significantly with the degree of I/R injury, as reflected by ALT levels. End-ischemic ex situ NMP results in activation of fibrinolysis, but not of coagulation. Markers of fibrinolysis activation correlate significantly with markers of I/R injury. High concentrations of D-dimer early after start of NMP can be considered a marker of severe I/R injury and a predictor of poor liver graft function.

Abstract TP8: A Novel and Thermally Efficient System for Focal Cerebral Hypothermia

Background: Focal cerebral hypothermia via intracarotid cold saline infusion (CSI) is an emerging therapy for acute stroke. Systems of cold saline delivery tested in humans have been thermally inefficient and failed to achieve target cortical temperatures. We applied a set of thermodynamic enhancements to improve thermal efficiency of CSI systems. Methods: We replicated the previously described CSI system composed of a cold saline bag (4°C), connected to a 5-Fr diagnostic angiographic catheter immersed in a model of aortic counterflow at 37°C and infusing at a rate of 33 cc/min. Three adjustments were introduced to the system: catheter insulation via a shuttle catheter, higher catheter flow rate (50 cc/min) and external cooling of the system. Those were collectively labeled Cold Preservation via External Cooling, Insulation and Supplemented flow (Cold-PRECISE). We compared the standard system with Cold-PRECISE and with each adjustment separately. Primary outcome was temperatures collected at tubing outlet, catheter tip and the estimated perfusate temperature. We used ANCOVA to compare primary outcomes adjusting for potential confounders. Results: Individual system adjustments improved thermal efficiency at all primary endpoints (Table 1). Compared to standard CSI, insulation and higher flow rate reduced perfusate temperatures by 0.33 and 1.96°C respectively (p &lt;0.001). External cooling reduced tubing outlet by 8.95°C (p 0.03) but had no statistically significant impact on perfusate temperature. The collective implementation of system adjustments had a synergistic effect with catheter tip and perfusate temperatures of 19.57 ± 0.34 and 29.87 ± 0.13 vs 27.13 ± 0.13 and 34.43 ± 0.05 respectively (p &lt;0.001). Conclusion: Cold-PRECISE system preserved coldness of locally infused saline, achieving perfusate temperatures that are estimated to meet target cortical temperatures. The clinical efficacy of this system should be evaluated in future clinical trials.

Promise of Normothermia

Normothermic ex vivo machine perfusion (NMP) is a novel preservation modality that has been investigated as a tool to preserve and protect organs from extended criteria donors (ECDs). This review summarizes the latest clinical and experimental progress in this field and tries to answer questions such as what are the future implications of NMP, what are its therapeutic potentials, and what are the limitations associated with this technology? New emerging data from clinical trials with NMP devices have demonstrated the safety and feasibility of this technology as well as its ability to preserve allograft function during the preservation period. NMP provides potential solutions to limitations associated with the standard cold preservation modality. It maintains the allografts in a physiological state, prevents depletion of cellular energy sources, enables resuscitation and assessment of the ECD organ, and opens the door for future ex vivo therapeutic interventions. Hence, it should increase donor pool and positively impact transplant outcomes.

Preservação de córneas: Um breve histórico

The first successful corneal transplantation used tissue from a living donor. The preservation of corneas gained momentum only after Filatov showed that a cadaver´s eye, when stored at low temperature, could be a source of viable corneas for transplants. Three methods proved to be useful for this purpose: moist chamber, cryopreservation, and liquid medium storage. They were introduced in that order. The first provides cold storage of corneas in situ for up to 2 days after death. It is still useful in underdeveloped regions of our planet. Eye banks practically abandoned the second method due to its complexity. Currently, the most popular storage technique is the third. Depending on the composition of the liquid medium, it allows for cold or warm preservation of the cornea ex situ. This article describes the characteristics of the three storage systems in quasi-chronological sequence

A study on the digitization of supply chains in agriculture - an Indian experience

In the present day context of globalisation, changing information needs of the farmers, increasing pressure of population on the food security system, encouraging the developing economy like India to look for various alternatives in supply chain management and its digitization for its efficient and sustainable agricultural development. India is likely to be considered as the food basket to the world constituting 52% of total land under cultivation as compared to global average of 11%. It is also producing 134.5 MT of fruits and vegetables but due to inadequate cold storage and preservation facilities and improper supply chain infrastructure; there is enormous loss of wastages. Supply chains are principally concerned with the flow of products and information between supply chain member organizations procurement of materials, transformation of materials into finished products, and distribution of those products to end customers. Today’s information-driven, integrated supply chains are enabling organizations to reduce inventory and costs, add product value, extend resources, accelerate time to market, and retain customers. Information Technology has started its dent in certain rural livelihoods especially the farmers in developing countries like India. IT can also do wonders in empowering small and marginal farmers who are operating in a complex, diverse and risk prone environment, who have poor access to information, especially regarding the production systems, customers and markets. In India, the limiting factors for farmers wanting to maximize their farm incomes are poor market linkages, poor access to quality farm-inputs, services and technology, lack of information about Government resources, institutions and extension services. ICT systems have pivotal role to play in market led extension activities. ICT s can connect the producers with buyers to initiate and sustain long term, mutually beneficial and sustainable professional relationships.

CARDIOPROTECTIVE EFFECTS OF NECROSTATIN‑1S AND NECROSULFONAMIDE IN THE MODEL OF PROLONGED STATIC COLD STORAGE OF THE DONOR RAT HEART

Objective . To investigate the effects of necroptosis inhibitors — necrosulfonamide and necrostatin‑1s — on morphofunctional characteristics of the myocardium in the model of prolonged cold preservation of the donor rat heart. Design and methods . The study was performed on 27 Wistar rats. The animals were divided into the following groups: 1) control (n = 7), 2) dimethylsulfoxide (DMSO, n = 6), 3) necrostatin‑1s (n = 8), 4) necrosulfonamide (n = 6). All test compounds were administered intraperitoneally 1 hour before the start of the experiment. Histidine-tryptophan-ketoglutarate (HTK) solution cooled to 4 °C was used as a preservative agent. DMSO was used as a solvent for necrostatin‑1s and necrosulfonamide. During the experiment, left ventricular pressure, heart rate and coronary flow were recorded. At the end of 8‑hour of ischemia and 2‑hour of reperfusion, myocardial infarct size was planimetrically evaluated. Results . Necrostatin-1s and necrosulfonamide show cardioprotective effects, as evidenced by the significantly lower levels of myocardial necrosis in these groups, as well as the best indices of intracardiac hemodynamics during reperfusion. In the groups of necrosulfonamide and necrostatin‑1s, myocardial infarct size was 32,2 ± 9,6 and 29,0 ± 9,2 %, respectively, which is significantly lower than in control groups and DMSO (54,4 ± 6,6 and 59,2 ± 5,6 %, respectively, p < 0,05). During reperfusion in necrosulfonamide and necrostatin‑1s groups, higher values of pulse intraventricular pressure and coronary flow rates were recorded, as well as lower diastolic intraventricular pressure, compared to control group and DMSO (p < 0,05). Conclusions . Necroptosis inhibitors necrostatin‑1s and necrosulfonamide improve morphofunctional characteristics of the myocardium state during prolonged cold preservation of the donor heart.

Nitrite enhances liver graft protection against cold ischemia reperfusion injury through a NOS independent pathway

ABSTRACTIntroduction: Nitrite has been found to protect liver graft from cold preservation injury. However, the cell signaling pathway involved in this protection remains unclear. Here, we attempt to clarify if the NOS pathway by using the NOS inhibitor, L-NAME (L-NG-Nitroarginine methyl ester).Animals and methods: Rat livers were conserved for 24 h at 4°C in (IGL-1) solution enriched or not with nitrite at 50 nM. In a third group, rats were pretreated with 50 mg/kg of L-NAME before their liver procurement and preservation in IGL-1 supplemented with nitrite (50 nM) and L-NAME (1 mM). After 24 h of cold storage, rat livers were ex-vivo perfused at 37°C during 2 h. Control livers were perfused without cold storage.Results: Nitrite effectively protected the rat liver grafts from the onset of cold I/R injury. L-NAME treatment did not abolish the beneficial effects of nitrite. Liver damage, protein oxidation and lipid peroxidation remained at low levels in both nitrite-treated groups when compared to IGL-1 group. Antioxidant enzyme activities and functional parameters were unchanged after NOS inhibition.Conclusion: Despite NOS inhibition by L-NAME, nitrite can still provide hepatic protection during cold I/R preservation. This suggests that nitrite acts through a NOS-independent pathway.

BacMam Delivery of a Protective Gene to Reduce Renal Ischemia-Reperfusion Injury.

Ischemia-reperfusion (I/R) injury remains the primary contributor to delayed graft function in kidney transplantation. The beneficial application of manganese superoxide dismutase (sod), delivered by a BacMam vector, against renal I/R injury has not been evaluated previously. Therefore, this study overexpressed sod-2 in proximal tubular epithelial (HK-2) cells and porcine kidney organs during simulated renal I/R injury. Incubation of HK-2 cells with antimycin A and 2-deoxyglucose resulted in a significant decrease in intracellular adenosine triphosphate (ATP) levels; following reperfusion, ATP levels significantly increased over time in cells overexpressing sod-2. In addition, lactate dehydrogenase (LDH) release declined over 72 h in BacMam-transduced injured cells. Ex vivo delivery of sod-2 significantly increased ATP levels in organs after 24 h of cold perfusion. In vitro and ex vivo results suggested that BacMam transduction successfully delivered sod-2, which reduced injury associated with I/R, by improving ATP cell content and decreasing LDH release with a subsequent increase in kidney tissue viability. These data provide further evidence for the potential application of BacMam as a gene delivery system for attenuating injury after cold preservation.

Lung inflation with hydrogen sulfide during the warm ischemia phase ameliorates injury in rat donor lungs via metabolic inhibition after cardiac death

Hydrogen sulfide attenuates lung ischemia-reperfusion injury when inhaled or administered intraperitoneally. This study investigated the effects of lung inflation with H2S during the warm ischemia phase on lung grafts from rat donors after cardiac death. One hour after cardiac death, donor lungs were inflated in situ for 2h with either O2 or H2S (O2 or H2S group) during the warm ischemia phase or were deflated as a control procedure (n=8). After 3h of cold preservation, lung transplantation was performed. During the warm ischemia phase, the metabolism and mitochondrial structures of donor lungs were analyzed. Arterial blood gas analysis was performed on the recipients. Protein expression in the graft of nuclear factor E2-related factor (Nrf)2 and nuclear factor kappa B (NF-κB) was analyzed by Western blotting, and static compliance, inflammation,oxidative stress, and cell apoptosis were assessed after 3h of reperfusion. When the O2 and H2S groups were compared with the control group, the mitochondrial structures were improved, and lactic acid levels, inflammation, oxidative stress, and cell apoptosis were significantly decreased; and glucose levels, as well as graft oxygenation and static compliance were increased. Simultaneously, the above indices showed further improvements, and the Nrf2 protein expression was significantly greater, and NF-κB protein expression was less in the H2S group than the O2 group. Lung inflation with H2S during the warm ischemia phase inhibited metabolism in donor lungs via mitochondrial protection, attenuated graft ischemic-reperfusion injury, and improved graft function through NF-κB-dependent anti-inflammatory and Nrf2-dependent antioxidative and antiapoptotic effects.