A brief review of the literature on traumatic anuria (acute tubular necrosis, lower nephron nephrosis) has been presented, including a complete bibliography. Special attention was paid to the pathology and pathogenesis of the syndrome and it was concluded that Oliver's recent work (271) probably comes closest to presenting the true picture. He describes tubular necrotic lesions for which the chemical toxins (mercuric chloride, carbon tetrachloride) were responsible, and tubulorhectic lesions which were characteristic of the shock kidney. These lesions could appear at any level in the renal tubule and were characterized by destruction of the basement membrane. Pigment casts were apparent if intravascular pigment release was associated with the illness. The work of Phillips, Van Slyke and associates (291, 292, 355, 356), of Oliver (271) and of Block et al (41) lead one to conclude that renal ischemia is the chief pathogenetic mechanism, though it is obvious that specific extrinsic renal toxins play a major role in specific cases. The role of hemoglobin appears to be chiefly in the production of obstructive casts later in the course of the disease; these pigments are precipitated in the lower nephron where urine is concentrated; and acidified, and dehydration and oliguria contribute to their formation. Three hundred rats were studied in eighteen experiments concerning crush syndrome. It was concluded that the most important single factor tending to aggravate the renal effects of crushing injury is the antecedent state of dehydration. Myoglobin is not an essential factor in the development of renal damage but tends to aggravate the existing uremia. Acute renal failure was seen to be a late effect of shock; animals developed acute tubular necrosis only if initial shock was severe, but not severe enough to produce death from circulatory failure. Development of this delicate balance of factors was aided by reduction of renal reserve by unilateral nephrectomy. A seldom described but distinct and consistent phenomenon was observed in the development of marked, immediate and persistent diuresis in response to the trauma of limb ligation. This polyuria was of a dilute urine and was taken as an indication of initial increased glomerular filtration followed by decreased reabsorption of water because of tubular damage. It was not an indication of a recovery phase as is recorded in the clinical syndrome. Testosterone propionate, desoxycorticosterone acetate, cortisone acetate and Compound F did not appear to be promising as therapeutic agents, although in one experiment Compound F showed some promise. Neither did combined therapy with testosterone and cortisone reduce the mortality rate or decrease uremia. Although there was no doubt that the syndrome of acute renal failure due to acute tubular necrosis could be produced in large numbers of these relatively inexpensive laboratory animals by dehydration and limb ligation, production could not altogether be standardized and the syndrome ran such a short course that serial observations were difficult to obtain and separation of shock deaths was occasionally impossible. It is felt that future work might well make use of some other laboratory animal, perhaps the dog or cat, and that an initial stress of controlled hypotension or renal artery occlusion could be used. It is also our opinion that further investigation into the value of Compound F as a therapeutic agent in this syndrome is justified. / Medicine, Faculty of / Graduate

Cyclosporin A (CsA) was incubated at concentrations of 5.0 x 10⁻⁶ M for 72 hours, and at concentrations of 1.0 and 0.5 x 10⁻⁶ M for 30 days with kangaroo rat proximal tubular epithelial cells (PtK₂) in order to evaluate its effects on the cytoskeleton. Alterations in the cytoskeleton were assessed by indirect immunofluorescence of viable cells, and by two dimensional electrophoresis of a high salt extract from the cells. There is a selective alteration of the cytokeratin intermediate filament organization in both the short term (5 x 10⁻⁶ M, 72 hr) and long term (1 and 0.5 x 10⁻⁶ M, 30 days) exposures. There are either peri-nuclear rings formed or the formation of a single aggregate clump of the cytokeratins within the cytoplasm. Other components of the cytoskeleton, the microtubules and the microfilaments remain unaffected at both short term and long term exposures. Along with this cytokeratin alteration in CsA exposed cells is the decrease or elimination of an acidic triplet of cytokeratin protein monomers, human equivalent K15 (50 kd), K16 (48 kd), K17 (46 kd). This may be related to CsA-associated nephrotoxicity.

This dissertation experimentally explores digital optical computing and optical interconnects with theoretical supports, from the physics of materials and the optimization of devices to system realization. The trend of optical computing is highlighted with the emphasis on the current development of its basic constituent elements, and a couple of algorithms selected to pave the way for utilizing bistable devices for their optical implementations. Optical bistable devices function as "optical transistors" in optical computing. The physics of dispersive optical bistability is briefly described. Bistable ZnS interference filters are discussed in detail regarding their linear and nonlienar characteristics. The optimization of switching characteristics for a bistable ZnS interference filter is discussed, and experimental results are shown. Symbolic substitution which fully takes advantage of regular optical interconnects constitutes two steps: pattern recognition and symbol scription. Two experiments on two digital pattern recognitions and one on a simple but complete symbolic substitution have been demonstrated. The extension of these experiments is an implementation of a binary adder. A one-bit full adder which is a basic block for a computer has been explored experimentally and demonstrated in an all-optical way. The utilization of a bistable device as a nonlinear decision-making element is further demonstrated in an associative memory experiment by incorporating a Vander Lugt matched filter to discriminate two partial fingerprints. The thresholding function of a bistable device enhances the S/N ratio and helps discrimination in associative memory. As the clocking speed of a computer goes higher, e.g. greater than several GHz, the clock signal distribution and packaging become serious problems in VLSI technology. The use of optical interconnects introduces a possible solution. A unique element for holographic optical interconnects, which combines advantages of computer generated hologram and DCG recording material, is discussed. Pattern design of a specific computer generated hologram and a proposed fabrication process are described. Experimental results suggest that this unique element has the capability of being tailored to perform multiple fan-out with resulting uniform tightly-focussed spots, and coupling between devices, e.g. source-to-fiber and fiber-to-waveguides, etc.

The critical function of the kidney is to regulate the body’s extracellular fluid volume to maintain homeostasis. When insults to the kidney occur, as in the case of kidney ischemia, the function of the kidney to filter metabolic wastes and reabsorb essential solutes is compromised, leading to a variety of clinical manifestations. Current metrics of kidney function are measured by the rise of a single analyte, the serum creatinine, which implies injury of the kidney tubule and its epithelial cells and is encapsulated by the term Acute Kidney Injury (AKI). Yet, creatinine does not specify the etiology, the cell type, or the molecular pathways that are affected by the acute decreases in kidney excretory function. During my thesis work, I hypothesized that there is a pathogenetic heterogeneity of kidney injury and a specificity of location, timing, and molecular mechanisms, unique to each of these three injury models: kidney ischemia, volume depletion, and urinary tract infection. Using genetic mouse models, RNA-sequencing, and a range of molecular biology techniques, I have found (1) kidney ischemia activates inflammatory responses, signal transduction pathways, and epithelial repair and reprogramming, that are not activated in volume depletion, (2) which in contrast, is a transient metabolic condition, inducing genes regulating energy metabolism that were reversible upon rehydration. Lastly, (3) I have found that urinary tract infection, particularly one that invades the kidney, involves a novel heme transport system in the collecting duct of the kidney, that may contribute to nutritional defenses against bacterial pathogens. Each of these findings is explored in specific aims and experiments, which I detail here in my thesis.

Reperfusion injury may mediate renal dysfunction following ischaemia. A murine model was developed to investigate the role of the tissue factor-thrombin-protease activated receptor pathway in renal ischaemia reperfusion injury (IRI). In this model, mice received 25 minutes of ischaemia and subsequent periods of reperfusion. C57BL6, protease activated receptor-1 (PAR-1) knockout mice, and tissue factor (TF) deficient mice were used. Following 24 hours IRI, PAR-1 deficiency resulted in protection against severe renal failure compared to the C57BL6 mice (creatinine, 118.2 ?? 6.3 vs 203 ?? 12 ??mol/l, p&lt0.001). This was confirmed by lesser tubular injury. By 48 hours IRI, this resulted in a survival benefit (survival, 87.5% vs 0%, p&lt0.001). Treatment of C57BL6 mice with hirudin, a specific thrombin inhibitor, offered renoprotection at 24 hours IRI (creatinine, 107 ?? 10 ??mol/l, p&lt0.001), leading to a 60% survival rate at 48 hours IRI (p&lt0.001). TF deficient mice expressing less than 1% of C57BL6 mouse TF were also protected (creatinine, 113.6 ?? 7 ??mol/l, p&lt0.001), with a survival benefit of 75% (p&lt0.001). The PAR-1 knockout, hirudin treated C57BL6 and TF deficient mice had reduced myeloperoxidase activity and tissue neutrophil counts compared to the C57BL6 mice, along with reduced KC and MIP-2 chemokine mRNA and protein expression. Hirudin treatment of PAR-1 knockout mice had no additional benefit over PAR-1 absence alone, suggesting no further contribution by activation of other protease activated receptors (creatinine at 24 hours IRI, 106.5 ?? 10.5 ??mol/l, p&gt0.05). Furthermore, immunofluoresence staining for fibrin(ogen) showed no difference between C57BL6 and PAR-1 knockout mice, suggesting no major contribution by fibrin in this model. Renal IRI resulted in increased levels of TF mRNA expression in the C57BL6, PAR-1 knockout, and hirudin treated C57BL6 mice compared to normal controls, suggesting that TF mRNA expression was upregulated in this model. This resulted in increased TF functional activity in the C57BL6 and PAR-1 knockout mice, but TF activity was negligible in hirudin treated C57BL6 and TF deficient mice. The data therefore suggests that the TF-thrombin cascade contributes to renal IRI by signalling via PAR-1 that then regulates chemokine gene expression and subsequent neutrophil recruitment.

Reperfusion injury may mediate renal dysfunction following ischaemia. A murine model was developed to investigate the role of the tissue factor-thrombin-protease activated receptor pathway in renal ischaemia reperfusion injury (IRI). In this model, mice received 25 minutes of ischaemia and subsequent periods of reperfusion. C57BL6, protease activated receptor-1 (PAR-1) knockout mice, and tissue factor (TF) deficient mice were used. Following 24 hours IRI, PAR-1 deficiency resulted in protection against severe renal failure compared to the C57BL6 mice (creatinine, 118.2 ?? 6.3 vs 203 ?? 12 ??mol/l, p&lt0.001). This was confirmed by lesser tubular injury. By 48 hours IRI, this resulted in a survival benefit (survival, 87.5% vs 0%, p&lt0.001). Treatment of C57BL6 mice with hirudin, a specific thrombin inhibitor, offered renoprotection at 24 hours IRI (creatinine, 107 ?? 10 ??mol/l, p&lt0.001), leading to a 60% survival rate at 48 hours IRI (p&lt0.001). TF deficient mice expressing less than 1% of C57BL6 mouse TF were also protected (creatinine, 113.6 ?? 7 ??mol/l, p&lt0.001), with a survival benefit of 75% (p&lt0.001). The PAR-1 knockout, hirudin treated C57BL6 and TF deficient mice had reduced myeloperoxidase activity and tissue neutrophil counts compared to the C57BL6 mice, along with reduced KC and MIP-2 chemokine mRNA and protein expression. Hirudin treatment of PAR-1 knockout mice had no additional benefit over PAR-1 absence alone, suggesting no further contribution by activation of other protease activated receptors (creatinine at 24 hours IRI, 106.5 ?? 10.5 ??mol/l, p&gt0.05). Furthermore, immunofluoresence staining for fibrin(ogen) showed no difference between C57BL6 and PAR-1 knockout mice, suggesting no major contribution by fibrin in this model. Renal IRI resulted in increased levels of TF mRNA expression in the C57BL6, PAR-1 knockout, and hirudin treated C57BL6 mice compared to normal controls, suggesting that TF mRNA expression was upregulated in this model. This resulted in increased TF functional activity in the C57BL6 and PAR-1 knockout mice, but TF activity was negligible in hirudin treated C57BL6 and TF deficient mice. The data therefore suggests that the TF-thrombin cascade contributes to renal IRI by signalling via PAR-1 that then regulates chemokine gene expression and subsequent neutrophil recruitment.

Indiana University-Purdue University Indianapolis (IUPUI) / The young field of gene therapy offers the promises of significant progress towards the treatment of many different types of human diseases. Gene therapy has been proposed as an innovative way to treat Acute Kidney Injury (AKI). Through proteomic analysis, the upregulation of two enzymes, IDH2 and SULT1C2, within the mitochondrial fraction has been identified following ischemic preconditioning, a treatment by which rat kidneys are protected from ischemia. Using the hydrodynamic fluid gene delivery technique, we were able to upregulate the expression of IDH2 and SULT1C2 in the kidney. We found that the delivery of IDH2 plasmid through hydrodynamic fluid delivery to the kidney resulted in increased mitochondrial oxygen respiration compared with injured kidneys without gene delivery. We also found that renal ischemic preconditioning altered the membrane fluidity of mitochondria. In conclusion, our study supports the idea that upregulated expression of IDH2 in mitochondria can protect the kidney against AKI, while the protective function of upregulated SULT1C2 needs to be further studied.

Indiana University-Purdue University Indianapolis (IUPUI) / Acute Kidney Injury (AKI) is a prevalent and significant problem whose primary treatment is supportive care. Ischemic preconditioning is a strategy used to protect organs from ischemic injury via a prior injury. Ischemic preconditioning in the kidneys has been shown to confer protection onto kidneys from subsequent ischemic insults with attenuated serum creatinine values in treated rats. In the preconditioned kidneys, the enzyme IDH2 was discovered to be upregulated in the mitochondria. Hydrodynamic fluid delivery to the kidney was found to be a viable technique for delivering this gene to the kidney, resulting in artificially upregulated expression of IDH2. Via a two-pronged effort to discern the functional significance of ischemic preconditioning and hydrodynamic IDH2 fluid injections, we performed mitochondrial oxygen respiration assays on both preconditioned and injected kidneys. We found that renal ischemic preconditioning resulted in no significant difference between sham and preconditioned, subsequently injured kidneys, which is similar to the results from the serum creatinine studies. Hydrodynamically IDH2-injected, and subsequently injured kidneys respire significantly better than vehicle injected, and subsequently injured kidneys, which shows that hydrodynamic injections of IDH2 protects kidneys against injury, and partially mimics the effects of preconditioning.

Indiana University-Purdue University Indianapolis (IUPUI) / Advancements in human genomics have simultaneously enhanced our basic understanding of the human body and ability to combat debilitating diseases. Historically, research has shown that there have been many hindrances to realizing this medicinal revolution. One hindrance, with particular regard to the kidney, has been our inability to effectively and routinely delivery genes to various loci, without inducing significant injury. However, we have recently developed a method using hydrodynamic fluid delivery that has shown substantial promise in addressing aforesaid issues. We optimized our approach and designed a method that utilizes retrograde renal vein injections to facilitate widespread and persistent plasmid and adenoviral based transgene expression in rat kidneys. Exogenous gene expression extended throughout the cortex and medulla, lasting over 1 month within comparable expression profiles, in various renal cell types without considerably impacting normal organ function. As a proof of its utility we by attempted to prevent ischemic acute kidney injury (AKI), which is a leading cause of morbidity and mortality across among global populations, by altering the mitochondrial proteome. Specifically, our hydrodynamic delivery process facilitated an upregulated expression of mitochondrial enzymes that have been suggested to provide mediation from renal ischemic injury. Remarkably, this protein upregulation significantly enhanced mitochondrial membrane potential activity, comparable to that observed from ischemic preconditioning, and provided protection against moderate ischemia-reperfusion injury, based on serum creatinine and histology analyses. Strikingly, we also determined that hydrodynamic delivery of isotonic fluid alone, given as long as 24 hours after AKI is induced, is similarly capable of blunting the extent of injury. Altogether, these results indicate the development of novel and exciting platform for the future study and management of renal injury.