Graft arterial stenosis in kidney en bloc grafts from very small pediatric donors: incidence, timing, and role of ultrasound in screening.

In previous studies with different donor selection criteria and noncontemporary surgical techniques, graft arterial stenosis (GAS) has been reported to occur more frequently in adult recipients of pediatric en bloc renal allografts (EBKT) as compared to single adult donor allografts. The purpose of our study was to evaluate the incidence of GAS within our EBKT recipient population and to evaluate clinical and imaging features of those cases with GAS. In a retrospective cohort study, we analyzed 182 EBKT performed at a single institution. We identified cases of suspected GAS based on clinical factors, lab results, and noninvasive imaging. Diagnosis of GAS was confirmed by digital subtraction angiography. Two EBKT recipients (1.1% of 182) had angiographically confirmed GAS at 2.5 and 4.5 months after transplant. In both cases, the stenoses were short segment within the proximal (perianastomotic) donor aorta, color Doppler ultrasound demonstrated peak systolic velocities of >400 cm/s, and poststenotic parvus tardus waveforms were present. Both patients underwent angioplasty and demonstrated postintervention improvement in renal function and blood pressure. Restenosis did not occur during follow up. In conclusion, recipients of EBKT have a low incidence of GAS, similar to the lowest reported for adult single allografts.

Optimizing recovery, utilization and transplantation outcomes for kidneys from small, ≤20 kg, pediatric donors.

The optimal balance between maximizing the number versus the outcome of transplantation utilizing kidneys from small (≤20 kg) pediatric donors remains unclear, complicated by the choice of single versus en bloc transplantation with their attendant technical risks. Using the Organ Procurement and Transplantation Network (OPTN) database, we examined kidney recovery and utilization patterns, and 1-year transplant outcomes by single kilogram weight strata. Between January 1, 2005 and June 30, 2010, 2352 kidneys from ≤20 kg donors were transplanted into 1531 recipients, 710 single kidney transplants (SKTs) and 821 en bloc kidney transplants (EBKTs). Increased donor weight was associated with higher rates of recovery, transplantation and SKT. Low donor weight (linear p < 0.001; quadratic p = 0.003), SKT versus EBKT (p = 0.008), increased cold ischemia time (p = 0.003), local versus nonlocal donor (p = 0.0044), low versus high volume center (p = 0.003) and the interaction term between center volume and donor weight (p = 0.0024) were associated with graft failure. Notably, lower donor weight exacerbated the negative impact of low center volume but did not worsen the negative impact of SKT on outcomes. Our data show that EBKT offers superior 1-year survival at the expense of accomplishing one rather than two transplants. However, SKTs yield excellent outcomes when performed at experienced centers.

Low progesterone concentration during the development of the first follicular wave reduces pregnancy per insemination of lactating dairy cows.

Objectives were to determine the effect of progesterone (P4) concentration on fertility of lactating dairy cows induced to ovulate follicles of the first follicular wave. Lactating dairy cows (n=989) at 38±3d postpartum were balanced by parity and body condition score and randomly assigned to 3 treatments: first follicular wave (FFW), first follicular wave with exogenous P4 (FFWP), or second follicular wave (SFW). All cows had their estrous cycle presynchronized with 2 injections of prostaglandin (PG) F(2α) given 14 d apart. Cows in the FFW and FFWP treatments started the ovulation synchronization protocol 3 d after the last PGF(2α) of the presynchronization protocol, whereas SFW cows received a GnRH injection (100 µg of gonadorelin diacetate; Cystorelin, Merial Ltd., Duluth, GA) 3 d after the last PGF(2α) of the presynchronization protocol and started the synchronization protocol 7 d later. The synchronization protocol consisted of GnRH on d -10, PGF(2α) on d -3, and GnRH concurrent with timed artificial insemination (AI) on d 0. Cows in the FFWP treatment received 2 controlled internal drug release inserts containing 1.38 g of P4 from d -8 to -3. Progesterone concentration was determined on d -10, -8, -6, -3, and 0 from all cows and at 7, 14, and 21 d after AI from a subsample of cows (n=170). Cows (n=715) had their ovaries scanned by ultrasound on d -10, -3, and 7 d. Pregnancy was diagnosed at 38 and 66 d after AI. Concentration of P4 from study d -8 to -3 was lowest for FFW cows (1.4±0.1 ng/mL) and similar between SFW (3.7±0.2 ng/mL) and FFWP (3.7±0.1 ng/mL) cows. Diameter of the dominant follicle on study d -3 was greater for FFW cows (16.5±0.3 mm) than for SFW cows (15.4±0.3 mm), but diameter of the dominant follicle of FFWP cows was not different (15.9±0.3 mm) compared with that of SFW and FFW cows. The incidence of multiple ovulation was largest for FFW cows (SFW=19.5, FFW=33.6, FFWP=19.0%), but pregnancy per AI (P/AI) at 66 d was smallest for FFW cows (SFW=38.9, FFW=22.3, FFWP=32.0%). Anovular cows in the SFW (19.4 vs. 42.8%) and FFWP (22.1 vs. 37.2%) treatments had reduced P/AI compared with cyclic cows, despite having similar or greater P4 concentration from study d -8 to -3, respectively. Estrus and ovulation synchronization protocols for lactating dairy cows must result in growth of ovulatory follicle under P4 concentration >2 ng/mL to ensure high P/AI.

Histopathologic Findings on Implantation Renal Allograft Biopsies Correlate With Kidney Donor Profile Index and 30-Day Serum Creatinine.

BACKGROUND: The Kidney Donor Profile Index (KDPI) provides a numerical estimate of deceased donor kidney quality. The KDPI uses 10 donor factors but it does not consider histopathologic findings. We examined whether the KDPI and its component donor factors correlate with the degree of histopathologic changes seen in implantation renal allograft biopsies. METHODS: All deceased donor kidney transplants at our institution from July 1, 2016 to March 15, 2017 that had an implantation biopsy were included. The biopsies were graded based on the Banff criteria for interstitial fibrosis, tubular atrophy, arterial intimal fibrosis, and arteriolar hyalinosis, as well as percent glomerulosclerosis. Linear and logistic regression were used to assess the correlation between histopathologic findings and KDPI and the ability of these variables to predict 30-day serum creatinine (SCr) and delayed graft function (DGF). RESULTS: One hundred thirty-four recipients from 107 donors were included. All histopathologic features examined correlated significantly with KDPI, with arteriolar hyalinosis correlating most strongly. Arteriolar hyalinosis was also associated with the most component donor factors of the KDPI. Histopathologic findings alone or in combination with KDPI predicted 30-day SCr but not DGF. Using the KDPI in combination with degree of interstitial fibrosis and tubular atrophy was the best predictor of 30-day SCr. CONCLUSION: Histopathologic changes seen in implantation renal allograft biopsies correlate with KDPI and predict 30-day SCr. Using a combination of donor histopathologic findings and KDPI may be the best predictor of short-term graft function.

Predictive value of pretransplant inflammatory markers in renal allograft survival and rejection in children.

Pretransplant (pre-Tx) inflammation has been associated with acute rejection (AR) in adult Tx recipients. Our study was performed to determine whether a single pre-Tx serum C-reactive protein (CRP), Neopterin (Neo), and IL-12 determination could predict outcome in pediatric renal Tx recipients. Pre-Tx sera from 51 children transplanted between 1985 and 2000 were analyzed for serum CRP, Neo, and IL-12 for correlation with Tx-related variables. Endpoints were graft loss and AR. Kaplan-Meier and log-rank statistics were used to compare rejection-free and overall graft survival at different quartiles for each marker. Cox regression analysis was performed to determine the independent effects of various pre-Tx variables on the endpoints. The mean age of the children at Tx was 11 years. The mean CRP, Neo, and IL-12 were 1.3 mg/L, 1.78 ng/mL and 123 pg/mL, respectively. At last-follow-up (mean 4.9 years after Tx), 50% of the children had experienced AR and 29% had lost their grafts. The mean CRP, Neo, and IL-12 were not different between the patients with versus without AR or graft loss (P > .4 for all). Neither rejection-free survival nor graft survival was affected by CRP, Neo, or IL-12 quartiles (log-rank test). Cox regression analysis demonstrated no predictive value of any marker on the outcomes. Unlike adults, a single pre-Tx determination of inflammatory markers was not predictive of AR or graft loss in children. The pathogenesis of AR may be different in children with a lesser contribution of alloantigen-independent factors such as chronic infections.

Augmentation of donor-specific transfusion and cyclosporine effects with dietary linoleic acid.

Increased prostaglandin production is a possible mechanism for the immunosuppressive effects of both cyclosporine and blood transfusions. Therefore, dietary supplementation with linoleic acid, a prostaglandin precursor, combined with either modality could act synergistically. Intraabdominal cardiac allografts were performed from Buffalo rat donors to Lewis recipients. Transplant recipients received a single donor-specific transfusion, low-dose cyclosporine (CsA, 1 mg/kd/d x 7 days), dietary supplementation with linoleic acid (LA, 16% of total calories) or a combination of the three modalities. CsA, DST or LA alone significantly prolonged allograft survival. Both CsA and LA acted synergistically with DST in further prolongation of survival--however, animals receiving all three modalities achieved 100% long-term survival. Augmentation of transfusion- and cyclosporine-induced immunosuppression with dietary prostaglandin precursor is possible.

Dietary immunoregulation of transfusion-induced immunosuppression.

Dietary supplementation with fatty acids was carried out to examine whether the type and timing of dietary manipulation would have an effect on transfusion-induced immunosuppression in a rat cardiac transplant model. Linoleic acid (LA), oleic acid (OA), and fish oil (FO) were used because of their different effects on arachidonic acid (AA) metabolism. Pretransplant inhibition of AA metabolism (OA) shortened graft survival when compared with water-fed controls. Posttransplant LA (AA precursor) significantly prolonged graft survival. Pre- and posttransplant supplementation of LA and OA resulted in even more prolongation and shortening of graft survival, respectively. These findings suggest that transfusion-induced immunosuppression is partially mediated by AA metabolites, which are necessary for the pretransplant induction and posttransplant maintenance of the suppressed state. Dietary immunoregulation of transfusion induced immunosuppression is possible. The timing of dietary intervention and type of lipid supplementation is important in regulation of the immune response.

Improvements in diabetic microangiopathy after successful simultaneous pancreas-kidney transplantation: a computer-assisted intravital microscopy study on the conjunctival microcirculation.

A computer-assisted intravital microscopy technology has been developed to noninvasively and objectively study diabetic microangiopathy in the conjunctival microcirculation of type-1 diabetics. Quantitative characterization of the conjunctival microcirculation was performed on 12 patients pre- and 18 months postsimultaneous pancreas-kidney transplantation (SPK). Healthy nondiabetic volunteers (n=12), solitary kidney (K) transplanted type-1 diabetics (n=5), and nontransplanted type-1 diabetics (n=12) served as controls. Pre-SPK diabetics showed abnormal-sized venules (diameter=66+/-7 microm) and reduced presence of arterioles (arteriole length/area=18+/-6 microm(-1)) compared with nondiabetic controls (53+/-4 microm; 31+/-8 microm(-1); P<0.05). The computed vascular perfusion capacity of the conjunctival microvasculature was diminished in the same patients (pre-SPK diabetics=49+/-9%; nondiabetic healthy controls=71+/-6%; P<0.05). Significant improvement in microangiopathy was observed in all post-SPK diabetics (diameter=58+/-6 microm; arteriole length/area=26+/-9 microm(-1); vascular perfusion=63+/-8%; P<0.05) 18 months post-SPK. Blood flow velocities in the conjunctival microcirculation in the same post-SPK patients showed noticeable but not significant improvements (nondiabetic controls=2.94+/-0.57 mm/sec; pre-SPK=1.23+/-0.49 mm/sec; post-SPK=1.65+/-0.42 mm/sec). The solitary kidney transplant controls (post-K) showed no significant improvements in diabetic microangiopathy, confirming the unique role of the pancreas in SPK. In general, significant improvements (P<0.05) in diabetic microangiopathy were observed in all 12 diabetics 18 months post-SPK but not in the controls.

Immunoregulation of transfusion-induced immunosuppression with inhibitors of the arachidonic acid metabolism.

To further define the role of arachidonic acid (AA) metabolites in transfusion-induced immunosuppression (TII), the effects of pharmacological manipulation of AA metabolism were examined in a rodent model. If the prostaglandins of the E series are mediators of TII, as has been recently hypothesized, then inhibition of cyclooxygenase (indomethacin) should abrogate whereas inhibition of lipoxygenase (nordihydroguaiaretic acid [NDGA]), or thromboxane synthetase (4-63557A) could potentiate the transfusion effect. Lewis rats received donor-specific transfusions from Buffalo rats in conjunction with one of the above inhibitors. Two weeks later they received intraabdominal Buffalo heart allografts or were used for one-way mixed lymphocyte reactions. Cyclooxygenase inhibition partially abrogated TII with shortened cardiac allograft survival. Lipoxygenase inhibition augmented TII, with depression of MLR and prolongation of allograft survival. Thromboxane synthetase inhibition had no effect. These results indicate that AA metabolites play a role in TII, and that immunoregulation via pharmacological manipulation of AA metabolism is possible.

Portal venous transfusion up-regulates Kupffer cell cyclooxygenase activity: a mechanism of immunosuppression in organ transplantation.

Portal venous transfusions (PVTs) of blood have been shown to induce significant immunosuppression in animal models of organ transplantation. A proposed mechanism of PVT-induced immunosuppression is via alteration of Kupffer cell arachidonic acid metabolism with increased secretion of the suppressive metabolite prostaglandin E2 (PGE2). This study assessed the hypothesis that PVT increases Kupffer cell PGE2 production via up-regulation of Kupffer cell phospholipase A2 (PLA2) as well as constitutive (COX1) and inducible (COX2) cyclooxygenase. Kupffer cells from Lewis rats were harvested 1 hr after PVT with either 1 ml of Wistar-Furth blood, systemic transfusion (SVT), or saline via portal vein (PVSal). After lipopolysaccharide stimulation, 24-hr Kupffer cell supernatant fractions were assayed for PGE2. PGE2 was increased after SVT (1465+/-234 pg/ml) compared with PVSal (597+/-99; P<0.01). PVT increased Kupffer cell PGE2 (5370+/-533; P<0.001 vs. SVT and vs. PVSal) even more substantially. Kupffer cells from PVT-treated rats were then cultured in the presence of inhibitors of PLA2, COX1, or COX2. When Kupffer cells were treated with mepacrine to inhibit PLA2 (5575+/-453 pg/ml), PGE2 production was not different from that by PVSal-treated controls (6467+/-614 pg/ml), but when Kupffer cells were incubated in the presence of the COX1 inhibitor flurbiprofen (3512+/-407 pg/ml) or the COX2 inhibitor nimesulide (2800+/-830 pg/ml), production was decreased 46.7% and 56.7%, respectively, over control activity without added inhibitor. PVT also increased Kupffer cells COX1 and COX2 mRNA as measured by Northern blot. Heart transplants were then performed from Wistar-Furth donors into Lewis recipients at the time of PVT, SVT, PVSal, or PVT + indomethacin (COX1/2 inhibitor). PVT prolonged allograft survival (12.0+/-0.9 days) compared with PVSal (6.3+/-0.3; P<0.01) or SVT (6.3+/-0.3; P<0.04). Indomethacin shortened graft survival when given with PVT (6.5+/-0.3 days). In summary, PVT increased Kupffer cell PGE2 production, up-regulated transcription of Kupffer cells COX1 and COX2 mRNA, and prolonged cardiac allograft survival. COX1/2 inhibition abrogated the effect of PVT. The results indicated that the immunosuppressive effect of PVT may be mediated by up-regulation of Kupffer cell COX1 and COX2. Manipulation of Kupffer cell arachidonic acid metabolism may be useful in augmentation of PVT-induced immunosuppression.

Selective targeting of Kupffer cells with liposomal butyrate augments portal venous transfusion-induced immunosuppression.

BACKGROUND: Enhanced Kupffer cell production of the immunosuppressive arachidonic acid metabolite prostaglandin E2 (PGE2) has been shown to be a mechanism of the immunosuppressive effect of portal venous transfusions (PVT). Butyrate, a four-carbon short-chain fatty acid, has received increased attention because of its ability to enhance gene transcription. This study tested the hypothesis that the intrahepatic delivery of butyrate enhances Kupffer cell PGE2 production and thus augments the immunosuppressive effect of PVT. METHODS: Butyrate was incorporated into liposomes and administered intravenously to Lewis rats. Control rats were administered liposomes without butyrate. Twenty-four hours after liposome injection, rats were administered a PVT of 1 ml of Wistar-Furth blood. Kupffer cells were isolated, and PGE2 and tumor necrosis factor-alpha levels were measured in the culture medium after 24 hr. Additionally, Kupffer cells from butyrate-treated and control animals were added to one-way mixed lymphocyte reaction cultures. RESULTS: Intrahepatic delivery of butyrate via liposomes increased Kupffer cell PGE2 (3800+/-1220 vs. 1010+/-119 pg/ml, P<0.05) and decreased tumor necrosis factor-alpha (1670+/-81 vs. 3360+/-415 pg/ml, P<0.01) production as compared with controls. Butyrate also augmented the Kupffer cell-mediated immunosuppression as demonstrated by significant depression of the mixed lymphocyte reaction (690+/-119 vs. 3850+/-148 cpm, P<0.01). CONCLUSION: The results support the hypothesis that intrahepatic delivery of butyrate enhances Kupffer cell PGE2 production, and specific targeting of Kupffer cells with liposomes containing immunomodulating agents such as butyrate may be a useful means of augmenting immunosuppression protocols in organ transplantation.

Pretransplant systemic inflammation and acute rejection after renal transplantation.

BACKGROUND: There are presently no established pre-transplant tests that consistently identify patients who may be at increased risk for acute rejection episodes after renal transplantation. We studied whether pretransplant serum levels of C-reactive protein (CRP), a marker for the presence of systemic inflammation, would predict the occurrence of acute rejection episodes after renal transplantation. METHODS: Pretransplant serum was tested for CRP level in 97 consecutive renal transplant recipients. Time to acute rejection after transplantation was stratified by CRP level and compared using the Kaplan-Meier method. In addition, Cox regression multivariate analysis was performed to assess whether any pretransplant covariates could independently predict the subsequent occurrence of acute rejection episodes. RESULTS: Pretransplant mean CRP levels were higher in patients who subsequently had a rejection episode versus those who had no rejection (22.2+/-2.9 vs. 11.7+/-1.8 microg/ml, respectively, P=0.003). Patients less than the median CRP value had a significantly longer time to rejection compared to those with higher CRP levels (P=0.002). Similarly, patients within the lowest CRP quartile had longer times to rejection when compared with the highest quartile (P=0.006). Cox proportional hazards regression multivariate analysis identified CRP level as the only independent pretransplant risk factor for rejection identified (P=0.044). CONCLUSIONS: Pretransplant systemic inflammation as manifested by elevated serum CRP level independently predicts the risk of acute rejection after renal transplantation and may be useful in stratifying patients at the time of transplantation according to immunological risk. Thus, assessment of pretransplant systemic inflammatory status may be helpful in prospective individualization of immunosuppression therapy after renal transplantation.

Increased transplantation of kidneys with multiple renal arteries in the laparoscopic live donor nephrectomy era: surgical technique and surgical and nonsurgical donor and recipient outcomes.

BACKGROUND: For anatomical and technical reasons, many transplant centers restrict laparoscopic live donor nephrectomy (in contrast with open live donor nephrectomy) to left kidneys. HYPOTHESIS: This change in surgical practice increases procurement and transplantation rates of live donor kidneys with multiple renal arteries (RAs), without affecting donor and recipient outcomes. DESIGN AND SETTING: Retrospective review at an academic tertiary care referral center comparing laparoscopically procured single vs multiple-RA kidney grafts (April 1997 to October 2000). PATIENTS: Seventy-nine consecutive left laparoscopic live kidney donors and 78 transplant recipients. MAIN OUTCOME MEASURES: Donor and recipient complications and postoperative length of stay; cold and warm ischemia time; operating time; short-term and long-term graft function; and survival. RESULTS: We noted multiple RAs in 21 (27%) of all kidneys. The proportion of donors with 1 or more perioperative complications was 19% in the single-RA group vs 10% in the multiple-RA group (P was not significant). For the recipients, we noted no significant differences between groups with respect to surgical complications, quality of early and late graft function, rejection rates, graft losses (all immunologic), and graft survival. Cold and warm ischemia time and length of stay were similar for donors and recipients in both groups. Median operating times were significantly longer for the multiple-RA vs single-RA group (difference, 41 minutes for donors and 45 minutes for recipients; P<.02). CONCLUSIONS: While the introduction of laparoscopic live donor nephrectomy has significantly increased the number of grafts with multiple RAs (compared with historical open controls), this change in practice is safe for both donors and recipients from a patient outcome-based perspective. However, from an economic perspective, the longer operating time associated with multiple-RA grafts provides strong added rationale for optimization of surgical instruments and techniques to make right-sided laparoscopic nephrectomy a routine intervention.

Evidence for intermolecular interaction as a necessary step for pore-formation activity and toxicity of Bacillus thuringiensis Cry1Ab toxin.

Based on the observation of large conductance states formed by Bacillus thuringiensis Cry toxins in synthetic planar lipid bilayers and the estimation of a pore size of 10-20 A, it has been proposed that the pore could be formed by an oligomer containing four to six Cry toxin monomers. However, there is a lack of information regarding the insertion of Cry toxins into the membrane and oligomer formation. Here we provide direct evidence showing that the intermolecular interaction between Cry1Ab toxin monomers is a necessary step for pore formation and toxicity. Two Cry1Ab mutant proteins affected in different steps of their mode of action (F371A in receptor binding and H168F in pore formation) were affected in toxicity against Manduca sexta larvae. Binding analysis showed that F371A protein bound more efficiently to M. sexta brush border membrane vesicles when mixed with H168F in a one to one ratio. These mutant proteins also recovered pore-formation activity, measured with a fluorescent dye with isolated brush border membrane vesicles, and toxicity against M. sexta larvae when mixed, showing that monomers affected in different steps of their mode of action can form functional hetero-oligomers.

Investigating fusion plasma instabilities in the Mega Amp Spherical Tokamak using mega electron volt proton emissions (invited).

The proton detector (PD) measures 3 MeV proton yield distributions from deuterium-deuterium fusion reactions within the Mega Amp Spherical Tokamak (MAST). The PD's compact four-channel system of collimated and individually oriented silicon detectors probes different regions of the plasma, detecting protons (with gyro radii large enough to be unconfined) leaving the plasma on curved trajectories during neutral beam injection. From first PD data obtained during plasma operation in 2013, proton production rates (up to several hundred kHz and 1 ms time resolution) during sawtooth events were compared to the corresponding MAST neutron camera data. Fitted proton emission profiles in the poloidal plane demonstrate the capabilities of this new system.