66 research outputs found
Isolation and in vitro cultivation of adrenal cells from mice
The adrenal gland consists of two tissues, cortex and medulla, united under one capsule. Adrenal stem/progenitor cells play a key role in development and homeostasis. Here, we describe a protocol for generating primary cultures of adrenal cells from mice. We describe techniques for separating the cortex and medulla, generating spheroid cultures containing stem- and progenitor cells, and for the differentiation into steroidogenic and chromaffin cells, respectively. This protocol enables analysis of various treatments before, during, or after differentiation. For complete details on the use and execution of this protocol, please refer to Rubin de Celis et al. (2015), Steenblock et al. (2018), and Werdermann et al. (2021)
Adrenal Hormone Interactions and Metabolism: A Single Sample Multi-Omics Approach
The adrenal gland is important for many physiological and pathophysiological processes, but studies are often restricted by limited availability of sample material. Improved methods for sample preparation are needed to facilitate analyses of multiple classes of adrenal metabolites and macromolecules in a single sample. A procedure was developed for preparation of chromaffin cells, mouse adrenals, and human chromaffin tumors that allows for multi-omics analyses of different metabolites and preservation of native proteins. To evaluate the new procedure, aliquots of samples were also prepared using conventional procedures. Metabolites were analyzed by liquid-chromatography with mass spectrometry or electrochemical detection. Metabolite contents of chromaffin cells and tissues analyzed with the new procedure were similar or even higher than with conventional methods. Catecholamine contents were comparable between both procedures. The TCA cycle metabolites, cis-aconitate, isocitate, and α-ketoglutarate were detected at higher concentrations in cells, while in tumor tissue only isocitrate and potentially fumarate were measured at higher contents. In contrast, in a broad untargeted metabolomics approach, a methanol-based preparation procedure of adrenals led to a 1.3-fold higher number of detected metabolites. The established procedure also allows for simultaneous investigation of adrenal hormones and related enzyme activities as well as proteins within a single sample. This novel multi-omics approach not only minimizes the amount of sample required and overcomes problems associated with tissue heterogeneity, but also provides a more complete picture of adrenal function and intra-adrenal interactions than previously possible
Transplantation of porcine adrenal spheroids for the treatment of adrenal insufficiency
Primary adrenal insufficiency is a life-threatening disorder, which requires lifelong hormone replacement therapy. Transplantation of xenogeneic adrenal cells is a potential alternative approach for the treatment of adrenal insufficiency. For a successful outcome of this replacement therapy, transplanted cells should provide adequate hormone secretion and respond to adrenal physiological stimuli. Here, we describe the generation and characterization of primary porcine adrenal spheroids capable of replacing the function of adrenal glands in vivo. Cells within the spheroids morphologically resembled adult adrenocortical cells and synthesized and secreted adrenal steroid hormones in a regulated manner. Moreover, the embedding of the spheroids in alginate led to the formation of cellular elongations of steroidogenic cells migrating centripetally towards the inner part of the slab, similar to zona Fasciculata cells in the intact organ. Finally, transplantation of adrenal spheroids in adrenalectomized SCID mice reversed the adrenal insufficiency phenotype, which significantly improved animals' survival. Overall, such adrenal models could be employed for disease modeling and drug testing, and represent the first step toward potential clinical trials in the future
Innovative multidimensional models in a high-throughput-format for different cell types of endocrine origin
The adrenal gland provides an important function by integrating neuronal, immune, vascular, metabolic and endocrine signals under a common organ capsule. It is the central organ of the stress response system and has been implicated in numerous stress-related disorders. While for other diseases, regeneration of healthy organ tissue has been aimed at such approaches are lacking for endocrine diseases - with the exception of type-I-diabetes. Moreover, adrenal tumor formation is very common, however, appropriate high-throughput applications reflecting the high heterogeneity and furthermore relevant 3D-structures in vitro are still widely lacking. Recently, we have initiated the development of standardized multidimensional models of a variety of endocrine cell/tissue sources in a new multiwell-format. Firstly, we confirmed common applicability for pancreatic pseudo-islets. Next, we translated applicability for spheroid establishment to adrenocortical cell lines as well as patient material to establish spheroids from malignant, but also benign adrenal tumors. We aimed furthermore at the development of bovine derived healthy adrenal organoids and were able to establish steroidogenic active organoids containing both, cells of cortical and medullary origin. Overall, we hope to open new avenues for basic research, endocrine cancer and adrenal tissue-replacement-therapies as we demonstrate potential for innovative mechanistic insights and personalized medicine in endocrine (tumor)-biology
Clinical improvement of Long-COVID is associated with reduction in autoantibodies, lipids, and inflammation following therapeutic apheresis
In the aftermath of the COVID-19 pandemic, we are witnessing an unprecedented wave of post-infectious complications. Most prominently, millions of patients with Long-Covid complain about chronic fatigue and severe post-exertional malaise. Therapeutic apheresis has been suggested as an efficient treatment option for alleviating and mitigating symptoms in this desperate group of patients. However, little is known about the mechanisms and biomarkers correlating with treatment outcomes. Here, we have analyzed in different cohorts of Long-Covid patients specific biomarkers before and after therapeutic apheresis. In patients that reported a significant improvement following two cycles of therapeutic apheresis, there was a significant reduction in neurotransmitter autoantibodies, lipids, and inflammatory markers. Furthermore, we observed a 70% reduction in fibrinogen, and following apheresis, erythrocyte rouleaux formation and fibrin fibers largely disappeared as demonstrated by dark field microscopy. This is the first study demonstrating a pattern of specific biomarkers with clinical symptoms in this patient group. It may therefore form the basis for a more objective monitoring and a clinical score for the treatment of Long-Covid and other postinfectious syndromes
Absence of Type I Interferon Autoantibodies or Significant Interferon Signature Alterations in Adults With Post-COVID-19 Syndrome
Genetic defects in the interferon (IFN) system or neutralizing autoantibodies against type I IFNs contribute to severe COVID-19. Such autoantibodies were proposed to affect post-COVID-19 syndrome (PCS), possibly causing persistent fatigue for >12 weeks after confirmed SARS-CoV-2 infection. In the current study, we investigated 128 patients with PCS, 21 survivors of severe COVID-19, and 38 individuals who were asymptomatic. We checked for autoantibodies against IFN-α, IFN-β, and IFN-ω. Few patients with PCS had autoantibodies against IFNs but with no neutralizing activity, indicating a limited role of type I IFNs in PCS pathogenesis. In a subset consisting of 28 patients with PCS, we evaluated IFN-stimulated gene activity and showed that it did not correlate with fatigue. In conclusion, impairment of the type I IFN system is unlikely responsible for adult PCS
A Rat Model of Post-Traumatic Stress Syndrome Causes Phenotype-Associated Morphological Changes and Hypofunction of the Adrenal Gland
BACKGROUND
Rats exposed to chronic predator scent stress mimic the phenotype of complex post-traumatic stress disorder (PTSD) in humans, including altered adrenal morphology and function. High- and low-anxiety phenotypes have been described in rats exposed to predator scent stress (PSS). This study aimed to determine whether these high- and low-anxiety phenotypes correlate with changes in adrenal histomorphology and corticosteroid production.
METHODS
Rats were exposed to PSS for ten days. Thirty days later, the rats' anxiety index (AI) was assessed with an elevated plus-maze test. Based on differences in AI, the rats were segregated into low- (AI ≤ 0.8, n = 9) and high- (AI > 0.8, n = 10) anxiety phenotypes. Plasma corticosterone (CORT) concentrations were measured by ELISA. Adrenal CORT, desoxyCORT, and 11-dehydroCORT were measured by high-performance liquid chromatography. After staining with hematoxylin and eosin, adrenal histomorphometric changes were evaluated by measuring the thickness of the functional zones of the adrenal cortex.
RESULTS
Decreased plasma CORT concentrations, as well as decreased adrenal CORT, desoxyCORT and 11-dehydroCORT concentrations, were observed in high- but not in low-anxiety phenotypes. These decreases were associated with increases in AI. PSS led to a significant decrease in the thickness of the zona fasciculata and an increase in the thickness of the zona intermedia. The increase in the thickness of the zona intermedia was more pronounced in low-anxiety than in high-anxiety rats. A decrease in the adrenal capsule thickness was observed only in low-anxiety rats. The nucleus diameter of cells in the zona fasciculata of high-anxiety rats was significantly smaller than that of control or low-anxiety rats.
CONCLUSION
Phenotype-associated changes in adrenal function and histomorphology were observed in a rat model of complex post-traumatic stress disorder
A Rat Model of Post-Traumatic Stress Syndrome Causes Phenotype-Associated Morphological Changes and Hypofunction of the Adrenal Gland
Background: Rats exposed to chronic predator scent stress mimic the phenotype of complex
post-traumatic stress disorder (PTSD) in humans, including altered adrenal morphology and function.
High- and low-anxiety phenotypes have been described in rats exposed to predator scent stress
(PSS). This study aimed to determine whether these high- and low-anxiety phenotypes correlate with
changes in adrenal histomorphology and corticosteroid production. Methods: Rats were exposed
to PSS for ten days. Thirty days later, the rats’ anxiety index (AI) was assessed with an elevated
plus-maze test. Based on differences in AI, the rats were segregated into low- (AI ≤ 0.8, n = 9)
and high- (AI > 0.8, n = 10) anxiety phenotypes. Plasma corticosterone (CORT) concentrations
were measured by ELISA. Adrenal CORT, desoxyCORT, and 11-dehydroCORT were measured
by high-performance liquid chromatography. After staining with hematoxylin and eosin, adrenal
histomorphometric changes were evaluated by measuring the thickness of the functional zones of the
adrenal cortex. Results: Decreased plasma CORT concentrations, as well as decreased adrenal CORT,
desoxyCORT and 11-dehydroCORT concentrations, were observed in high- but not in low-anxietyphenotypes. These decreases were associated with increases in AI. PSS led to a significant decrease in
the thickness of the zona fasciculata and an increase in the thickness of the zona intermedia. The increase
in the thickness of the zona intermedia was more pronounced in low-anxiety than in high-anxiety
rats. A decrease in the adrenal capsule thickness was observed only in low-anxiety rats. The nucleus
diameter of cells in the zona fasciculata of high-anxiety rats was significantly smaller than that of
control or low-anxiety rats. Conclusion: Phenotype-associated changes in adrenal function and
histomorphology were observed in a rat model of complex post-traumatic stress disorder
Host factor PLAC8 is required for pancreas infection by SARS-CoV-2
Although mounting evidence demonstrated that pancreas is infected by SARS-CoV-2
the severity and pathophysiology of pancreatic COVID-19 disease are still unclear. Here
we investigated the consequences of SARS-CoV-2 infection of the pancreas and the role
of Placenta-associated protein-8 (PLAC8). Our data showed pancreatic damage in
patients who died from COVID-19. Notably, circulating pancreatic enzymes stratified
patients according to COVID-19 severity and outcome. PLAC8 expression was
associated with SARS-CoV-2 infection in postmortem analysis of COVID-19 patients and
functional assays demonstrated the requirement of PLAC8 in SARS-CoV-2 pancreatic
infection. Full SARS-CoV-2 infectious virus revealed a requirement of PLAC8 for efficient
viral infection of pancreatic cell lines. Finally, we observed colocalization of PLAC8 and
SARS-CoV-2 in the pancreas of deceased patients. In conclusion, our data confirm the
human pancreas as a SARS-CoV-2 target and demonstrate the requirement of PLAC8
for SARS-CoV-2 pancreatic infection thereby opening new target opportunities for
COVID-19-associated pancreatic pathogenesis.N
Adrenal cortical and chromaffin stem cells: Is there a common progeny related to stress adaptation?
The adrenal gland is a highly plastic organ with the capacity to adapt the body homeostasis to different physiological needs. The existence of stem-like cells in the adrenal cortex has been revealed in many studies. Recently, we identified and characterized in mice a pool of glia-like multipotent Nestin-expressing progenitor cells, which contributes to the plasticity of the adrenal medulla. In addition, we found that these Nestin progenitors are actively involved in the stress response by giving rise to chromaffin cells. Interestingly, we also observed a Nestin-GFP-positive cell population located under the adrenal capsule and scattered through the cortex. In this article, we discuss the possibility of a common progenitor giving rise to subpopulations of cells both in the adrenal cortex and medulla, the isolation and characterization of this progenitor as well as its clinical potential in transplantation therapies and in pathophysiology
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