Different methods of measuring testicular blood flow in the rat were compared in an attempt to find an accurate method for measuring physiological testicular blood flow. It was found that both the Xenon- 133 clearance technique and the radioactive microsphere technique probably reflect true physiological blood flow in the testis. The microsphere method was used to study some functional aspects of testicular blood flow. There was a significant positive correla­tion between the testicular blood flow and the outflow of testosterone in the spermatic vein, indicating that testicular hormone secretion may be affected indirectly via a primary effect on testicular blood flow. Intra-arterial infusion of LH caused a significant decrease in the vascular resistance of the testis. However, the effect was small in comparison with the simultaneous effect of LH on plasma testosterone concentration, indicating that blood flow changes are not critically involved in the acute effect of LH on testicular endocrine function. Infusion of epinephrine or norepinephrine did not induce any absolute changes in testicular blood flow, but norepinephrine caused an in­crease in testicular vascular resistance. Both catecholamines caused significant depressions in plasma testosterone concentration. It was concluded that the catecholamine induced reductions in testosterone concentration were not due to a vascular effect on the testis. Testicular blood flow and Leydig cell function in the cryptorchid and heated testis were also studied. There was a significant increase in relative blood flow in the cryptorchid testis, probably due to an highly altered morphology consisting of a relative increase in inter­stitial tissue containing blood vessels. Furthermore, it was found that the testosterone levels, in spermatic vein blood from the cryptor­chid testis, were highly reduced in comparison to the corresponding values for the scrotal testis and the outflow of testosterone from the cryptorchid testis was estimated to be only 13% of that from the scrotal one. This result suggested that the Leydig cell function was greatly impaired in the cryptorchid testis. The vasculature of the testis is relatively insensitive to local heating since no effects on vascular resistance were observed when warming the testis to ab­dominal temperature. On the other hand, there was a significant in­crease in blood flow in the testis at 41 and 43° C, which are tempera­tures known to induce cessation of spermatogenesis in the rat. The acute response of the testis to LH stimulation was reduced when warm­ing the scrotum to 41 and 43° C. This strongly indicates an impaired Leydig cell function at these temperatures. Since blood flow was in­creased at these temperatures it was concluded that the reduced Leydig cell responsiveness to LH was unrelated to testicular perfusion. / digitalisering@umu.se

No description available.

No description available.

Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disease which may affect multiple organ systems. Interferon alpha (IFNα) and autoantibodies that form immune complexes with nuclear antigens (ANA) are hallmarks believed to drive the disease into a vicious circle of inflammation, tissue damage, autoantigen exposure and autoantibody production. In SLE, the disease course is characterized by episodes of exacerbations alternating with remissions. In order to best treat the patient it is important to closely monitor symptoms and signs of disease activity. Because of the disease heterogeneity, no single biomarker has yet been found to reflect SLE disease activity in general, although antidouble stranded DNA (anti-dsDNA) antibodies sometimes indicate activity, primarily with renal involvement, and constitutes an item of the SLE disease activity score SLEDAI-2K. However, the method of anti-dsDNA measurement is not standardized and therefore varies between different laboratories. In many other inflammatory conditions, such as rheumatoid arthritis and during bacterial infections, the C-reactive protein (CRP) level is a good indicator of ongoing inflammation, but in SLE and during viral infections, CRP commonly fails to reflect the degree of inflammation. Both viral infections and SLE are characterized by IFNα, and we thus aimed to elucidate whether IFNα can inhibit CRP production. Further, four assays for anti-dsDNA antibody measurements were evaluated with regard to SLE disease specificity and activity, and a new potential biomarker of inflammation, the soluble urokinase plasminogen activator receptor (suPAR), was assessed in relation to disease activity and organ damage. An in vitro inhibitory effect of IFNα on CRP transcription and production was found in hepatocytes, and this was consolidated by in vivo studies of CRP and IFNα in sera from well-characterized SLE patients (KLURING; Kliniskt lupusregister i nordöstra Götaland). Here, CRP and disease activity were associated among patients without IFNα and without a CRP lowering gene variant (SNP rs1205). The poor disease activity compliance of CRP could therefore be explained, at least in part, by polymorphisms in the CRP gene and increased levels of IFNα. Critical differences between the methods measuring anti-dsDNA were found regarding disease specificity and ability to reflect disease activity and the results suggests the Crithidia luciliae immunofluorescence test (CLIFT) for diagnostic purposes and a bead-based multiplex assay (FIDIS) for monitoring of disease activity. Evaluation of suPAR in SLE revealed no association of suPAR with disease activity, but interestingly instead with accumulated organ damage. suPAR could therefore possibly be used to advert patients at high risk of organ damage. A detailed biological and clinical characterization of established and emerging SLE biomarkers is of importance since it may improve the clinical management as well as increase the knowledge about disease mechanisms.

Intellectual disability (ID) is a common neurodevelopmental condition, often caused by genetic defects. De novo variation (DNV) is an important cause of ID, especially in severe or syndromic forms of the disorder. Next generation sequencing has been a successful application for finding pathogenic variation in ID patients. The main focus of this thesis is to use whole exome sequencing (WES) and whole genome sequencing (WGS) to identify pathogenic variants in undiagnosed ID patients. In Paper I, WES was used in family trios to identify pathogenic DNVs in patients diagnosed with ID in combination with epilepsy. This work led to the identification of several DNVs in both new and known disease genes, including the first report of variation in the HECW2 gene in association with neurodevelopmental disorder and epilepsy. Paper II is the first independent validation of PIGG as a disease-causing gene in patients with developmental disorder. We used WES to identify the homozygous variation in PIGG, and transcriptome analysis as well as flow-cytometry studies were used to validate the pathogenicity of the PIGG variation. We discovered that PIGG variation give different effects in different cell types, contributing new insights into the disease mechanism. Paper III is also an application of WES in trio families with patients diagnosed with ID in order to identify causal variants, a strategy similar to that of Paper I. Several pathogenic variants were identified in this study; in particular, the gene NAA15 is highlighted as a new disease gene, and was recently confirmed in independent studies. This study also adds evidence to support that variation in the PUF60 gene is causing the symptoms in patients with Verheij syndrome. In Paper IV, WGS was used to analyze families with consanguineous marriages. All families in this study had been previously analyzed with WES without finding a disease cause. A number of new disease-causing variants were identified in the study, including a first validation of FRMD4A as a disease-associated gene. This study also shows that WGS performs better than WES in finding variants, even for variants in coding parts of the genome.

Sammanfattning Syfte och frågeställningar Syftet med studien var att undersöka graden och kvaliteten av informationen gällande kost och fysisk aktivitet som ges av sjukvården till patienter med diagnosen Ankyloserande spondylit. Vidare undersöktes om det fanns ett samband mellan graden av fysisk aktivitet samt kostval hos patienter med Ankyloserande Spondylit och mängden information som getts av sjukvården. Ytterligare undersöktes om det fanns samband mellan graden fysisk aktivitet samt kostval och hur de uppskattade sitt välmående. Metod Studien är en kvantitativ tvärsnittsstudie där populationen är individer diagnostiserade med Ankyloserande Spondylit. Urvalet har skett med hjälp av bekvämlighets- och tillgänglighetsurval. En elektronisk enkätundersökning delades ut till populationen med hjälp av sociala medier, totalt inkom 130 svar. Svaren på enkäten samlades in och kodades om till siffror för att kunna analyseras i SPSS. Analysmetoderna som användes var deskriptiv statistik, variansanalys, t-test samt korrelationssamband. Resultat 81 % av de som svarade hade fått information om fysisk aktivitet och kostval kopplat till sitt sjukdomstillstånd. Det var 42,5% som ansåg att de hade fått tillräcklig information och 57,5% som upplevde att det inte varit tillräckligt. Medelvärdet på hur mycket information de fått om kost var 2,8 och medelvärdet för kvaliteten på informationen var 2,9. I samma fråga fast gällande fysisk aktivitet var medelvärdet 7,4 och medelvärdet för den kvaliteten var 6,8. Det fanns en skillnad mellan hur mycket information de hade fått om fysisk aktivitet och hur många minuter högintensiv fysisk aktivitet de utförde per tillfälle (p = 0,009). Skillnader kunde hittas mellan hur mycket information de fått om kost och fiberintag (p = 0,03). De som hade skattat lågt på enkäten om sjukdomsförloppet (BASDAI) drack mindre alkohol per tillfälle (p=0,02). Skillnader kunde inte hittas mellan hur mycket information de fått om kost och livsmedel som exkluderats ur kosten. Slutsats Populationen i denna studie hade fått mycket information om fysisk aktivitet men inte om kost. Kvaliteten på informationen om fysisk aktivitet ansågs vara relativt hög och kvaliteten om kost låg. De som hade fått lite information om fysisk aktivitet var fler minuter högintensivt fysiskt aktiva. De som mådde bättre i sjukdomsförloppet drack mindre alkohol per tillfälle.

A growing body of results indicate that immunological alterations and oxidative stress are of importance in various mental disorders. The inflammatory changes are possible to detect both in plasma and cerebrospinal fluid (CSF), and different psychiatric disorder exhibit similar changes indicating a common underlying mechanism. The antioxidants are of importance to regulate the redox balance and control the inflammatory processes but the causal relationship between psychiatric disorders and increased oxidative stress is however not fully clarified. Two important antioxidants; taurine and glutathione (GSH), have been suggested to have central nervous system (CNS)-protective properties. They have been found to fluctuate in several mental disorders including schizophrenia and depression but the clinical relevance need further studies. The general aim of this thesis was to increase the understanding of taurine and the GSH in depression and schizophrenia, two major mental disorders, in comparison to healthy controls. Correlations between glutathione and taurine levels in blood and CSF were analyzed in healthy male volunteers and we identified a complex pattern of associations showing that the CSF concentration was influenced by body mass index (BMI), age, intraspinal pressure, plasma concentrations the previous day and possible genetic factors. Electroconvulsive therapy (ECT) is used in the treatment of severely depressed patients. In blood collected before the first and after the third ECT, we found a significant decrease in plasma taurine in patients responding to the treatment, while total glutathione was unaltered. In a group of olanzapine treated patients with schizophrenia or schizoaffective disorders, we analysed taurine and glutathione in plasma and CSF and compared with healthy male and female volunteers. We observed increased plasma taurine levels in patients compared with controls, but no difference in CSF taurine and no alteration in glutathione. This thesis indicates that taurine might play a role in mental disorders such as depression and schizophrenia. Increased knowledge about the complex regulation of taurine and glutathione might provide new insights into the impact of redox balance in the pathophysiology of psychiatric disorder and contribute to a future personalization of the treatment.

Lysosomes are acidic organelles that are critically involved in a number of physiological processes, including macromolecule degradation, endocytosis, autophagy, exocytosis and cholesterol homeostasis. Several pathological conditions, such as cancer, neurodegenerative disorders and lysosomal storage diseases, involve lysosomal disturbances, indicating the importance of the organelle for correct cellular function. The aim of this thesis was to investigate the role of lysosomes in cell death signaling. Previous studies have shown that permeabilization of the lysosomal membrane and release of hydrolytic enzymes such as cathepsin D to the cytosol occurs during apoptosis. We identified Bid and 14-3-3 proteins as cytosolic targets of cathepsin D in human fibroblasts. Truncated Bid, generated by cathepsin D proteolytic cleavage, stimulates Bax-mediated release of pro-apoptotic factors from the mitochondria, thereby engaging the intrinsic pathway to apoptosis. Since the presence of cathepsins in the cytosol is sufficient to induce apoptosis, the permeability of the lysosomal membrane influences the fate of the cell. In this thesis, we demonstrated that the stability of the lysosomal membrane can be manipulated by altering the lysosomal cholesterol content. Cells with high lysosomal cholesterol content were less prone to undergo apoptosis when challenged with stimuli known to induce lysosome-mediated cell death. In addition, cholesterol accumulation was associated with increased expression of lysosome-associated membrane proteins and storage of other lipids; however, these factors did not contribute to lysosomal stabilization. Lysosomal membrane permeabilization and cathepsins contribute to ultraviolet (UV) irradiation-induced apoptosis. We demonstrate plasma membrane damage induced by UVA irradiation to be rapidly repaired by lysosomal exocytosis in human keratinocytes. Despite efficient plasma membrane resealing, the cells underwent apoptosis, which was dependent on early activation of caspase-8. The activation of caspase-8 was lysosome-dependent and occurred in vesicles positive for lysosomal markers. This thesis demonstrates the importance of lysosomal stability for apoptosis regulation and that this stability can be influenced by drug intervention. Modulation of the lysosomal membrane permeability may have potential for use as a therapeutic strategy in conditions associated with accelerated or repressed apoptosis.

No description available.

Nanoparticles are small particles with a size range of 10-1000 nm. They exist all around us, in make-up, dust and even food. They can enter our bloodstream through different pathways such as inhalation and cause thrombosis and multiple organ failure. They can be modified to act as drug deliverers and can treat even hard to reach places because of their small size. Studies have shown that the activation of the coagulation system and complement system is dependent on the size of the nanoparticle. This study’s main focus was to determine if there was a difference in the degree of activation on hemostasis and innate immunity by using four different nanoparticle sizes. The Chandler loop model makes it possible for blood to incubate with the nanoparticles and still be circulating in 37oC similar to the situation in the body. ELISA was thereafter performed on the plasma to determine the concentration of thrombin- antithrombin complex (TAT), C3a and Terminal Complement Complex (TCC). The most activating particles size on the complements system was 260 nm and for the coagulation system it was the 75 nm. FXII assay was performed and the results collaborated with the findings from the ELISA that the smallest particle sizes are most activating on the coagulation system. Taken together, smaller nanoparticle sizes are activating the coagulation system while the bigger nanoparticle sizes are more activating on the complement system. To confirm these results additional research should be performed to statistically confirm the importance of these findings.