Biology subjects

Lee, J. H.

Publications and source records attributed to Lee, J. H..

16 recordsLinked to original sources

bioRxiv2018

Protect TUDCA stimulated CKD-derived hMSCs against the CKD-Ischemic disease via upregulation of PrPC

Although autologous human mesenchymal stem cells (hMSCs) are a promising source for regenerative stem cell therapy, the barriers associated with pathophysiological conditions in this disease limit therapeutic applicability to patients. We proved treatment of CKD-hMSCs with TUDCA enhanced the mitochondrial function of these cells and increased complex I & IV enzymatic activity, increasing PINK1 expression and decreasing mitochondrial O2*- and mitochondrial fusion in a PrPC-dependent pathway. Moreover, TH-1 cells enhanced viability when co-cultured in vitro with TUDCA-treated CKD-hMSC. In vivo, tail vein injection of TUDCA-treated CKD-hMSCs into the mouse model of CKD associated with hindlimb ischemia enhanced kidney recovery, the blood perfusion ratio, vessel formation, and prevented limb loss, and foot necrosis along with restored expression of PrPC in the blood serum of the mice. These data suggest that TUDCA-treated CKD-hMSCs are a promising new autologous stem cell therapeutic intervention that dually treats cardiovascular problems and CKD in patients.

cell biology↗

bioRxiv2018

Decoupling between causal understanding and awareness during learning and inference

Causal reasoning is a principal higher-cognitive ability of humans, however, much remains unknown, including (a) the type (systematic versus intermixed) and order (inductive-then-deductive or vice versa) of experience that best achieves causal-chain extraction; (b) how inferences generalize to novel problems, especially with one-shot experience; and (c) how metacognition, reflected in uncertainty of ones knowledge, relates to actual knowledge. We tested people on a realistic cancer biology task (e.g., seroc chemicals inducing tumors with subsequent effects). Systematic experience was superior, with some evidence that the inductive-then-deductive order promoted stronger one-shot generalization. Notably, uncertainty was decoupled from actual knowledge, with the deductive-then-inductive group being overconfident, likely reflecting lack of awareness of the inductive component; while those with successful one-shot generalization held lower confidence, reflecting generalization with minimal experience, while remaining skeptical. Our findings clarify processes underlying causal reasoning, and reveal a complex relationship between causal reasoning and metacognitive awareness of it.

neuroscience↗

bioRxiv2018

The tumor suppressor p53 promotes carcinoma invasion and collective cellular migration

SummaryLoss of function of the tumor suppressor p53 is generally thought to increase cell motility and invasiveness. Using 2-D confluent and 3-D spheroidal cell motility assays with bladder carcinoma cells and colorectal carcinoma cells, we report, to the contrary, that loss of p53 can decrease cell motility and invasion.\n\nAbstractFor migration of the single cell studied in isolation, loss of function of the tumor suppressor p53 is thought to increase cell motility. Here by contrast we used the 2-D confluent cell layer and the 3-D multicellular spheroid to investigate how p53 impacts dissemination and invasion of cellular collectives. We used two human carcinoma cell lines, the bladder carcinoma EJ and the colorectal carcinoma HCT116. We began by replicating single cell invasion in the traditional Boyden chamber assay, and found that the number of invading cells increased with loss of p53, as expected. In the confluent 2-D cell layer, however, for both EJ and HCT, speeds and effective diffusion coefficients for the p53 null types compared to their p53 expressing counterparts were significantly smaller. Compared to p53 expressers, p53 null cells exhibited more organized cortical actin rings together with reduced front-rear cell polarity. Furthermore, loss of p53 caused cells to exert smaller traction forces upon their substrates, and reduced formation of cryptic lamellipodia. In a 3-D collagen matrix, p53 consistently promoted invasion of the multicellular spheroids into surrounding matrix. Together, these results show that p53 expression in these carcinoma model systems increases collective cellular migration and invasion. As such, these studies point to paradoxical contributions of p53 in single cell versus collective cellular migration.

cancer biology↗

bioRxiv2018

Structural determinants and genetic modifications enhance BMP2 stability and extracellular secretion

The short half-life and use of recombinant bone morphogentic protein (BMP)-2 in large doses poses major limitations in the clinic. Events regulating post-translational processing and degradation of BMP2 in situ, linked to its secretion, have not been understood. Towards identifying mechanisms regulating intracellular BMP2 stability, we first discovered that inhibiting proteasomal degradation enhances both intracellular BMP2 level and its extracellular secretion. Next, we identified BMP2 degradation occurs through an ubiquitin-mediated mechanism. Since ubiquitination precedes proteasomal turnover and mainly occurs on lysine residues of nascent proteins, we systematically mutated individual lysine residues within BMP2 and tested them for enhanced stability. Results revealed that substitutions on four lysine residues within the pro-BMP2 region and three in the mature region increased both BMP2 turnover and extracellular secretion. Structural modeling revealed key lysine residues involved in proteasomal degradation occupy a lysine cluster near proprotein convertase cleavage site. Interestingly, mutations within these residues did not affect biological activity of BMP2. These data suggest preventing intracellular proteasomal loss of BMP2 through genetic modifications can overcome limitations related to its short half-life.

cell biology↗

bioRxiv2018

Selection of an Appropriate Empiric Antibiotic Regimen in Culture-Negative Hematogenous Vertebral Osteomyelitis

The aim of this study was to determine which antibiotic combinations are appropriate for culture-negative hematogenous vertebral osteomyelitis (HVO), based on the antibiotic-susceptibility pattern of organisms isolated from cases of culture-proven HVO. We conducted a retrospective chart review of adult patients with microbiologically proven HVO in five tertiary-care hospitals over a 7-year period. The appropriateness of empiric antibiotic regimens was assessed based on the antibiotic susceptibility profiles of isolated bacteria. In total, 358 cases of microbiologically proven HVO were identified. The main causative pathogens identified were methicillin-susceptible Staphylococcus aureus (33.5%), followed by methicillin-resistant S. aureus (MRSA) (24.9%), aerobic gram-negative bacteria (21.8%), and Streptococcus species (11.7%). Extended spectrum {beta}-lactamase (ESBL)-producing Enterobacteriaceae and anaerobes accounted for only 1.7% and 1.4%, respectively, of the causative pathogens. Based on the susceptibility results of isolated organisms, levofloxacin plus rifampicin was appropriate in 73.5%, levofloxacin plus clindamycin in 71.2%, and amoxicillin-clavulanate plus ciprofloxacin in 64.5% of cases. These oral combinations were more appropriate for treating community-acquired HVO (85.8%, 84.0%, and 80.4%, respectively) than healthcare-associated HVO (52.6%, 49.6%, and 37.6%, respectively). Vancomycin combined with ciprofloxacin, ceftriaxone, ceftazidime, or cefepime was similarly appropriate (susceptibility rates of 93.0%, 94.1%, 95.8%, and 95.8%, respectively). In conclusion, in a setting with a high prevalence of MRSA HVO, oral antibiotic combinations may be suboptimal for treatment of culture-negative HVO and should be used only in patients with community-acquired HVO. Vancomycin combined with fluoroquinolone or a broad-spectrum cephalosporin was appropriate in most cases of HVO in this study.

epidemiology↗

bioRxiv2018

Multiplexed single-cell RNA-seq via transient barcoding for drug screening

To simultaneously analyze multiple samples of various conditions with scRNA-seq, we developed a universal sample barcoding method through transient transfection of SBOs. A 48-plex drug treatment experiment of pooled samples analyzed by a single run of Drop-Seq revealed a unique transcriptome response for each drug and target-specific gene expression signatures at the single-cell level. Our cost-effective method is widely applicable for single-cell profiling of multiple experimental conditions.

genomics↗

bioRxiv2018

Exploring Mechanisms of Inhibition of Amyloid Seeding of Transthyretin

Amyloid deposition of the hormone transporter transthyretin causes familial and sporadic amyloidoses. The current treatment for familial cases is gene-therapy by liver transplantation. However, this procedure is often insufficient to stop subsequent cardiac deposition. Our recent work has shown that preformed amyloid fibrils present in the heart by the time of surgery can template or seed further polymerization of native transthyretin. No drugs have been approved to stop or slow this seeding process; the only treatment option is heart transplantation. Here we explore two potential inhibitory mechanisms. Of clinical significance, we found that tetramer stabilization does not hinder amyloid seeding. In contrast, binding of the peptide inhibitor TabFH2 to ex-vivo fibrils efficiently inhibits amyloid seeding in a tissue-independent manner. Our findings point to inhibition of amyloid seeding by peptide inhibitors as a potential therapeutic approach to be further explored.

biochemistry↗

bioRxiv2018

Temporal Evolution of Cortical Ensembles Promoting Remote Memory Retrieval

Studies of amnesic patients and animal models support a systems consolidation model, which posits that explicit memories formed in hippocampus are transferred to cortex over time1-6. Prelimbic cortex (PL), a subregion of the medial prefrontal cortex, is required for the expression of learned fear memories from hours after learning until weeks later7-12. While some studies suggested that prefrontal cortical neurons active during learning are required for memory retrieval13-15, others provided evidence for ongoing cortical circuit reorganization during memory consolidation10,16,17. It has been difficult to causally relate the activity of cortical neurons during learning or recent memory retrieval to their function in remote memory, in part due to a lack of tools18. Here we show that a new version of targeted recombination in active populations, TRAP2, has enhanced efficiency over the past version, providing brain-wide access to neurons activated by a particular experience. Using TRAP2, we accessed PL neurons activated during fear conditioning or 1-, 7-, or 14-day memory retrieval, and assessed their contributions to 28-day remote memory. We found that PL neurons TRAPed at later retrieval times were more likely to be reactivated during remote memory retrieval, and more effectively promoted remote memory retrieval. Furthermore, reducing PL activity during learning blunted the ability of TRAPed PL neurons to promote remote memory retrieval. Finally, a series of whole-brain analyses identified a set of cortical regions that were densely innervated by memory-TRAPed PL neurons and preferentially activated by PL neurons TRAPed during 14-day retrieval, and whose activity co-varied with PL and correlated with memory specificity. These findings support a model in which PL ensembles underlying remote memory undergo dynamic changes during the first two weeks after learning, which manifest as increased functional recruitment of cortical targets.

neuroscience↗

bioRxiv2018

Sparse recurrent excitatory connectivity in the cortical microcircuit of the adult mouse and human

Generating a comprehensive description of cortical networks requires a large-scale, systematic approach. To that end, the Allen Institute is engaged in a pipeline project using multipatch electrophysiology, supplemented with 2-photon optogenetics, to characterize connectivity and synaptic signaling between classes of neurons in adult mouse and human cortex. We focus on producing results detailed enough for the generation of computational models and enabling comparison with future studies. Here we report our examination of intralaminar connectivity within each of several classes of excitatory neurons. We find that connections are sparse but present among all excitatory cell types and layers we sampled, with the most sparse connections in layers 5 and 6. Almost all mouse synapses exhibited short-term depression with similar dynamics. Synaptic signaling between a subset of layer 2/3 neurons; however, exhibited facilitation. These results contribute to a body of evidence describing recurrent excitatory connectivity as a conserved feature of cortical microcircuits.

neuroscience↗

bioRxiv2018

Cancer exosomes induce tumor neo-neurogenesis potentiating tumor growth

Patients with densely innervated tumors do worse than those with less innervated cancers. We hypothesize that neural elements are acquired by a tumor-induced process, called neo-neurogenesis. Here, we use PC12 cells in a simple system to test this hypothesis. PC12 cells extend processes, called neurites, only when appropriately stimulated. Using this system, we show that patient tumors release vesicles (exosomes) which induce PC12 neurite outgrowth. Using a cancer mouse model, we show that tumor cells compromised in exosome release grow slower and are less innervated than controls indicating a contribution of innervation to disease progression. We find that neo-neurogenesis is mediated in part by the axonal guidance molecule, EphrinB1, contained in exosomes. These findings support testing EphrinB1 blockers to inhibit tumor innervation and improve survival.\n\nOne Sentence SummaryTumors release exosomes which not only promote their own innervation but also potentiate their growth.

cancer biology↗

bioRxiv2017

Biologically plausible mechanisms underlying motor response correction during reward-based decision-making

AO_SCPLOWBSTRACTC_SCPLOWOur propensity to acclimate to new surroundings and choose a goal-directed behavior for a maximal reward (i.e., optimal outcome) is natural, for it affects our survival. A line of studies suggested that anterior cingulate cortex (ACC) could be a potential hub for adaptive behaviors. For instance, an experimental study noted ACCs contribution to selecting motor responses for maximal rewards; it found 1) that ACC neurons were selectively activated when the reward was reduced and 2) that suppression of ACC activity impaired monkeys ability to change motor responses to obtain the maximal reward. To probe ACC functions in adaptive behaviors, we sought biologically-plausible mechanisms to account for the experimental findings mentioned above by utilizing a computational model. Our simulation results raise the possibility that ACC can correct behavioral responses by reading out and updating the motor plans (guiding future motor responses) stored in prefrontal cortex (PFC).

neuroscience↗

bioRxiv2017

Efficient in situ barcode sequencing using padlock probe-based BaristaSeq

Cellular DNA/RNA tags (barcodes) allow for multiplexed cell lineage tracing and neuronal projection mapping with cellular resolution. Conventional approaches to reading out cellular barcodes trade off spatial resolution with throughput. Bulk sequencing achieves high throughput but sacrifices spatial resolution, whereas manual cell picking has low throughput. In situ sequencing could potentially achieve both high spatial resolution and high throughput, but current in situ sequencing techniques are inefficient at reading out cellular barcodes. Here we describe BaristaSeq, an optimization of a targeted, padlock probe-based technique for in situ barcode sequencing compatible with Illumina sequencing chemistry. BaristaSeq results in a five-fold increase in amplification efficiency, with a sequencing accuracy of at least 97%. BaristaSeq could be used for barcode-assisted lineage tracing, and to map long-range neuronal projections.\n\nKey PointsO_LIIn situ sequencing by gap-filling padlock probes is limited by the strand displacement of DNA polymerases\nC_LIO_LIIllumina sequencing chemistry offers superior signal-to-noise ratio in situ compared to sequencing by ligation\nC_LIO_LIBaristaSeq as an accurate method for barcode sequencing in situ with improved gap-filling efficiency\nC_LI

molecular biology↗

bioRxiv2017

Accurate detection of low-level somatic mutations with technical replication for next-generation sequencing

Accurate genome-wide detection of somatic mutations with low variant allele frequency (VAF, <1%) has proven difficult, for which generalized, scalable methods are lacking. Herein, we describe a new computational method, called RePlow that we developed to detect low-VAF somatic mutations based on simple, library-level replicates for next-generation sequencing on any platform. Through joint analysis of replicates, RePlow is able to remove prevailing background errors in next-generation sequencing analysis, facilitating remarkable improvement in the detection accuracy for low-VAF somatic mutations (up to [~]99% reduction in false positives). The method was validated in independent cancer panel and brain tissue sequencing data. Our study suggests a new paradigm with which to exploit an overwhelming abundance of sequencing data for accurate variant detection.

genomics↗

bioRxiv2017

Cortical circuit based lossless neural integrator for perceptual decision-making

The intrinsic uncertainty of sensory information (i.e., evidence) does not necessarily deter an observer from making a reliable decision. Indeed, uncertainty can be reduced by integrating (accumulating) incoming sensory evidence. It is widely thought that this accumulation is instantiated via recurrent rate-code neural networks. Yet, these networks do not fully explain important aspects of perceptual decision-making, such as a subjects ability to retain accumulated evidence during temporal gaps in the sensory evidence. Here, we utilized computational models to show that cortical circuits can switch flexibly between retention and integration modes during perceptual decision-making. Further, we found that, depending on how the sensory evidence was readout, we could simulate stepping and ramping activity patterns, which may be analogous to those seen in different studies of decision-making in the primate parietal cortex. This finding may reconcile these previous empirical studies because it suggests these two activity patterns emerge from the same mechanism.

neuroscience↗

bioRxiv2017

Multiple reference genome sequences of hot pepper reveal the massive evolution of plant disease resistance genes by retroduplication

Transposable elements (TEs) provide major evolutionary forces leading to new genome structure and species diversification. However, the role of TEs in the expansion of disease resistance gene families has been unexplored in plants. Here, we report high-quality de novo genomes for two peppers (Capsicum baccatum and C. chinense) and an improved reference genome (C. annuum). Dynamic genome rearrangements involving translocations among chromosome 3, 5 and 9 were detected in comparison between C. baccatum and the two other peppers. The amplification of athila LTR-retrotransposons, members of the gypsy superfamily, led to genome expansion in C. baccatum. In-depth genome-wide comparison of genes and repeats unveiled that the copy numbers of NLRs were greatly increased by LTR-retrotransposon-mediated retroduplication. Moreover, retroduplicated NLRs exhibited great abundance across the angiosperms, with most cases lineage-specific and thus recent events. Our study revealed that retroduplication has played key roles in the emergence of new disease-resistance genes in plants.

plant biology↗

bioRxiv2016

Visual processing mode switching regulated by VIP cells

The responses of neurons in mouse primary visual cortex (V1) to visual stimuli depend on behavioral states. Specifically, surround suppression is reduced during locomotion. Although locomotion-induced vasoactive intestinal polypeptide positive (VIP) interneuron depolarization can account for the reduction of surround suppression, the functions of VIP cell depolarization are not fully understood. Here we utilize a firing rate model and a computational model to elucidate the potential functions of VIP cell depolarization during locomotion. Our analyses suggest 1) that surround suppression sharpens the visual responses in V1 to a stationary scene, 2) that depolarized VIP cells enhance V1 responses to moving objects by reducing self-induced surround suppression and 3) that during locomotion V1 neuron responses to some features of the moving objects can be selectively enhanced. Thus, VIP cells regulate surround suppression to allow pyramidal neurons to optimally encode visual information independent of behavioral state.

neuroscience↗