T-Cell Mediated Immune Rejection of Beta-2-Microglobulin Knockout Induced Pluripotent Stem Cell-Derived Kidney Organoids.

Immune evasive induced pluripotent stem cell (iPSC)-derived kidney organoids, known as "stealth" organoids, hold promise for clinical transplantation. To address immune rejection, we investigated the impact of genetically modifying human leukocyte antigen (HLA) class I in kidney organoids prior to transplantation. By using CRISPR-Cas9, we successfully knocked out beta-2-microglobulin (B2M), resulting in iPSCs devoid of HLA class I surface expression. In vitro, the B2M knockout protected kidney organoids derived from these iPSCs against T-cell rejection. To assess in vivo protection, unmodified (control) and B2M-/- kidney organoids were transplanted into humanized mice engrafted with human peripheral blood mononuclear cells (PBMCs). Successful engraftment of human PBMCs was confirmed, and after 4 weeks, we observed no discernible difference in the infiltration rate, proliferation, or cytotoxicity of CD4+ and CD8+ T cells between control and B2M-/- organoids. Both groups of organoids showed compromised tissue integrity, displaying tubulitis and loss of tubule integrity. Notably, while B2M-/- organoids failed to express HLA class I on their cell surface, there was preexisting expression of HLA class II in both control and B2M-/- organoids transplanted into mice with human PBMCs. HLA class II expression was not limited to antigen-presenting cells but also evident in epithelial cells of the kidney organoid, posing an additional immunological challenge to its transplantation. Consequently, we conclude that B2M knockout alone is insufficient to protect iPSC-derived kidney organoids from T-cell-mediated immune rejection. Additionally, our findings suggest that modulating HLA class II signaling will be necessary to prevent rejection following transplantation.

Motor sequence chunking is impaired by basal ganglia stroke.

Our main aim was to determine whether individuals with stroke that affected the basal ganglia, organized movement sequences into chunks in the same fashion as neurologically intact individuals. To address this question, we compared motor response times during the performance of repeated sequences that were learned, and thus may be planned in advance, with random sequences where there is minimal if any advance preparation or organization of responses. The pattern of responses illustrated that, after basal ganglia stroke, individuals do not chunk elements of the repeated sequence into functional sub-sequences of movement to the same extent as neurologically intact age-matched people. Limited chunking of learned movements after stroke may explain past findings that show overall slower responses even when sequences of action are learned by this population. Further, our data in combination with other work, suggest that chunking may be a function of the basal ganglia.

Manipulating time-to-plan alters patterns of brain activation during the Fitts' task.

Fitts' law predicts that there is an essential trade-off between speed and accuracy during movement. Past investigations of Fitts' law have not characterized whether advance planning of upcoming fast and accurate movements impacts either behavior or patterns of brain activation. With an event-related functional magnetic resonance imaging (fMRI) paradigm, we investigated the neural correlates of advance planning and movement difficulty of rapid, goal-directed aimed movements using a discrete version of the classic Fitts' task. Our behavioral data revealed strong differences in response time, initial movement velocity, and end-point accuracy based on manipulation of both time to plan movements and response difficulty. We discovered a modulation of the neural network associated with executing the Fitts' task that was dependent on the availability of time to plan the upcoming movement and motor difficulty. Specifically, when time to plan for the upcoming movement was available, medial frontal gyrus (BA 10), pre-SMA (BA 6), putamen and cerebellar lobule VI were uniquely active to plan movements. Further, their activation correlated with behavioral measures of movement. In contrast, manipulating movement difficulty invoked a different pattern of brain activations in regions that are known to participate in motor control, including supplementary motor area (BA 6), sensory motor cortex (BA 4, 3, 2) and putamen. Our finding that medial frontal gyrus (BA 10) was important for discrete, fast and accurate movements expands the known role of this brain region, which in the past has been identified as a cognitive processing system supporting stimulus-oriented attending. We now extend this conceptualization to include motor functions such as those employed for processing for rapid, goal-directed aimed movements.

Comparative morphological and growth kinetics studies of human hair bulb papilla cells and root sheath fibroblasts in vitro.

Dermal papillae isolated from anagen hair bulbs obtained from biopsy specimens from five subjects with normal hair pattern, and fibroblasts derived from the mesenchymal root sheaths (RSF) of the same hair follicles were separately grown in culture and the cell-cycle distribution pattern on different days was analyzed by applying DNA flow cytometry (FCM). Papilla cells (PC) exhibited distinctive morphological features by forming cell aggregates differing from RSF with respect to cell shape and growth pattern. They also proliferated remarkably more slowly than RSF. DNA-FCM analysis showed that both PC and RSF demonstrated synchronous fluctuations in the percentage of cells in G1/0, S and G2 + M phases during the period of subculture.

Effects of epidermal growth factor, fibroblast growth factor, minoxidil and hydrocortisone on growth kinetics in human hair bulb papilla cells and root sheath fibroblasts cultured in vitro.

Comparative studies on growth kinetics of cultivated human hair bulb papilla cells (PCs) and hair root sheath fibroblasts (RSFs) yielded evidence of some peculiarities of PCs in both proliferative behavior and morphological growth pattern. As the dermal papilla, essentially supporting the nutrition of matrix epithelium, can be considered a target tissue for agents influencing maintenance of hair growth, we studied the effects of epidermal growth factor (EGF), fibroblast growth factor (FGF), minoxidil (Mino), and hydrocortisone (HC) on the proliferation of PCs and RSFs, both gained from dissected hair follicles of scalp biopsy specimens of two male adults and separately cultured in vitro. EGF and FGF proved to increase proliferation of both PCs and RSFs most, yet at a different intensity for each cell group. HC slowed proliferation, and Mino failed to influence growth of PCs and RSFs.

[Tissue reactions with administration of piezoelectric shock waves in lithotripsy of salivary calculi]. / Gewebereaktionen unter Applikation von piezoelektrischen Stosswellen zur Lithotripsie von Speichelsteinen.

Before clinical application of an extracorporeal piezoelectric lithotripter to treat sialolithiasis, tissue reaction during shockwave application was examined in vitro and in experiments with animals. Application of shockwaves to human tissue in vitro showed neither macroscopic nor microscopic effects. In animal experiments, the acute experiment (16 rabbits, Chinchilla-Bastard) revealed minor bleeding in the parenchyma of the parotid gland, while the chronic experiment (14 rabbits, Chinichilla-Bastard) revealed no morphologic tissue damage to the parotid region of the rabbit, as a result of piezoelectric shockwaves. However, when the eye was placed in the shockwave focal area and the shockwaves were transmitted via the fissura orbitalis to the endocranium, brain damage could be detected morphologically. In conclusion, the authors feel that the clinical application of extracorporeal piezoelectric fragmentation of salivary stones is justified, provided that a reliable positioning of the patient and exact sonographic location of the concrement are possible.