Synaptic plasticity in human thalamocortical assembloids.

Synaptic plasticities, such as long-term potentiation (LTP) and depression (LTD), tune synaptic efficacy and are essential for learning and memory. Current studies of synaptic plasticity in humans are limited by a lack of adequate human models. Here, we modeled the thalamocortical system by fusing human induced pluripotent stem cell-derived thalamic and cortical organoids. Single-nucleus RNA-sequencing revealed that most cells in mature thalamic organoids were glutamatergic neurons. When fused to form thalamocortical assembloids, thalamic and cortical organoids formed reciprocal long-range axonal projections and reciprocal synapses detectable by light and electron microscopy, respectively. Using whole-cell patch-clamp electrophysiology and two-photon imaging, we characterized glutamatergic synaptic transmission. Thalamocortical and corticothalamic synapses displayed short-term plasticity analogous to that in animal models. LTP and LTD were reliably induced at both synapses; however, their mechanisms differed from those previously described in rodents. Thus, thalamocortical assembloids provide a model system for exploring synaptic plasticity in human circuits.

Selective chemogenetic inactivation of corticoaccumbal projections disrupts trait choice impulsivity.

Impulsive choice has enduring trait-like characteristics and is defined by preference for small immediate rewards over larger delayed ones. Importantly, it is a determining factor in the development and persistence of substance use disorder (SUD). Emerging evidence from human and animal studies suggests frontal cortical regions exert influence over striatal reward processing areas during decision-making in impulsive choice or delay discounting (DD) tasks. The goal of this study was to examine how these circuits are involved in decision-making in animals with defined trait impulsivity. To this end, we trained adolescent male rats to stable behavior on a DD procedure and then re-trained them in adulthood to assess trait-like, conserved impulsive choice across development. We then used chemogenetic tools to selectively and reversibly target corticostriatal projections during performance of the DD task. The prelimbic region of the medial prefrontal cortex (mPFC) was injected with a viral vector expressing inhibitory designer receptors exclusively activated by designer drugs (Gi-DREADD), and then mPFC projections to the nucleus accumbens core (NAc) were selectively suppressed by intra-NAc administration of the Gi-DREADD actuator clozapine-n-oxide (CNO). Inactivation of the mPFC-NAc projection elicited a robust increase in impulsive choice in rats with lower vs. higher baseline impulsivity. This demonstrates a fundamental role for mPFC afferents to the NAc during choice impulsivity and suggests that maladaptive hypofrontality may underlie decreased executive control in animals with higher levels of choice impulsivity. Results such as these may have important implications for the pathophysiology and treatment of impulse control, SUDs, and related psychiatric disorders.

Innate frequency-discrimination hyperacuity in Williams-Beuren syndrome mice.

Williams-Beuren syndrome (WBS) is a rare disorder caused by hemizygous microdeletion of ∼27 contiguous genes. Despite neurodevelopmental and cognitive deficits, individuals with WBS have spared or enhanced musical and auditory abilities, potentially offering an insight into the genetic basis of auditory perception. Here, we report that the mouse models of WBS have innately enhanced frequency-discrimination acuity and improved frequency coding in the auditory cortex (ACx). Chemogenetic rescue showed frequency-discrimination hyperacuity is caused by hyperexcitable interneurons in the ACx. Haploinsufficiency of one WBS gene, Gtf2ird1, replicated WBS phenotypes by downregulating the neuropeptide receptor VIPR1. VIPR1 is reduced in the ACx of individuals with WBS and in the cerebral organoids derived from human induced pluripotent stem cells with the WBS microdeletion. Vipr1 deletion or overexpression in ACx interneurons mimicked or reversed, respectively, the cellular and behavioral phenotypes of WBS mice. Thus, the Gtf2ird1-Vipr1 mechanism in ACx interneurons may underlie the superior auditory acuity in WBS.

Patterns in Tacrolimus Variability and Association with De Novo Donor-Specific Antibody Formation in Pediatric Kidney Transplant Recipients.

BACKGROUND AND OBJECTIVES: High tacrolimus intrapatient variability has been associated with inferior graft outcomes in patients with kidney transplants. We studied baseline patterns of tacrolimus intrapatient variability in pediatric patients with kidney transplants and examined these patterns in relation to C1q-binding de novo donor-specific antibodies. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: All tacrolimus levels in participants who underwent kidney-only transplantation at a single pediatric center from 2004 to 2018 (with at least 12-month follow-up, followed until 2019) were analyzed to determine baseline variability. Intrapatient variability was defined using the coefficient of variation (SD/mean ×100%) of all samples in a 6-month moving window. Routine de novo donor-specific antibody measurements were available for a subgroup of patients transplanted in 2010-2018. Cox proportional hazards models using tacrolimus intrapatient variability as a time-varying variable were used to examine the association between intrapatient variability and graft outcomes. The primary outcome of interest was C1q-binding de novo donor-specific antibody formation. RESULTS: Tacrolimus intrapatient variability developed a steady-state baseline of 30% at 10 months post-transplant in 426 patients with a combined 31,125 tacrolimus levels. Included in the outcomes study were 220 patients, of whom 51 developed C1q-binding de novo donor-specific antibodies. De novo donor-specific antibody formers had higher intrapatient variability, with a median of 38% (interquartile range, 28%-48%) compared with 28% (interquartile range, 20%-38%) for nondonor-specific antibody formers (P<0.001). Patients with high tacrolimus intrapatient variability (coefficient of variation >30%) had higher risk of de novo donor-specific antibody formation (hazard ratio, 5.35; 95% confidence interval, 2.45 to 11.68). Patients in the top quartile of tacrolimus intrapatient variability (coefficient of variation >41%) had the strongest association with C1q-binding de novo donor-specific antibody formation (hazard ratio, 11.81; 95% confidence interval, 4.76 to 29.27). CONCLUSIONS: High tacrolimus intrapatient variability was strongly associated with de novo donor-specific antibody formation.

Physiotherapists Both Reproduce and Resist Biomedical Dominance when Working With People With Low Back Pain: A Qualitative Study Towards New Praxis.

Despite recommendations to incorporate physical and psychosocial factors when providing care for people with back pain, research suggests that physiotherapists continue to focus on biological aspects. This study investigated how interpersonal and institutional norms influence this continued enactment of the biological aspects of management. We used theoretically-driven analysis, drawing from Foucauldian notions of power, to analyse 28 ethnographic observations of consultations and seven group discussions with physiotherapists. Analysis suggested that physiotherapy training established expectations of what a physiotherapist 'should' focus on, and institutional circumstances strongly drew the attention of physiotherapists towards biological aspects. Resistance to these forces was possible when, for example, physiotherapists reflected upon their practice, used silences and pauses during consultations, and actively collaborated with patients. These circumstances facilitated use of non-biomedical management approaches. Findings may assist physiotherapists to rework the enduring normative focus on biomedical aspects of care when providing care for patients with back pain.

Cortical control of striatal fast-spiking interneuron synchrony.

Inhibitory fast-spiking interneurons in the dorsal striatum regulate actions and action strategies, including habits. Fast-spiking interneurons are widely believed to synchronize their firing due to the electrical synapses formed between these neurons. However, neuronal modelling data suggest convergent cortical input may also drive synchrony in fast-spiking interneuron networks. To better understand how fast-spiking interneuron synchrony arises, we performed dual whole-cell patch clamp electrophysiology experiments to inform a simple Bayesian network modelling cortico-fast-spiking interneuron circuitry. Dual whole-cell patch clamp electrophysiology revealed that while responsivity to corticostriatal input activation was high in fast-spiking interneurons, few of these neurons exhibited electrical coupling in adult mice. In simulations of a cortico-fast-spiking interneuron network informed by these data, the degree of glutamatergic cortical convergence onto fast-spiking interneurons significantly increased fast-spiking interneuron synchronization while manipulations of electrical coupling between these neurons exerted relatively little impact. These results suggest that the primary source of functional coordination of fast-spiking interneuron activity in adulthood arises from convergent corticostriatal input activation. KEY POINTS: Electrical synapses between striatal fast-spiking interneurons in adult mice occur in ∼8% of assayed pairs. Coincident, convergent cortical input onto fast-spiking interneurons significantly contributes to fast-spiking interneuron synchrony. Electrical synapses between fast-spiking interneurons provide only minor enhancement of fast-spiking interneuron synchrony. These results suggest a mechanism by which adult mouse fast-spiking interneurons of the striatum synchronize in the face of declining expression of the electrical synapse-forming connexin-36 protein.

Alzheimer's disease-associated U1 snRNP splicing dysfunction causes neuronal hyperexcitability and cognitive impairment.

Recent proteome and transcriptome profiling of Alzheimer's disease (AD) brains reveals RNA splicing dysfunction and U1 small nuclear ribonucleoprotein (snRNP) pathology containing U1-70K and its N-terminal 40-KDa fragment (N40K). Here we present a causative role of U1 snRNP dysfunction to neurodegeneration in primary neurons and transgenic mice (N40K-Tg), in which N40K expression exerts a dominant-negative effect to downregulate full-length U1-70K. N40K-Tg recapitulates N40K insolubility, erroneous splicing events, neuronal degeneration and cognitive impairment. Specifically, N40K-Tg shows the reduction of GABAergic synapse components (e.g., the GABA receptor subunit of GABRA2), and concomitant postsynaptic hyperexcitability that is rescued by a GABA receptor agonist. Crossing of N40K-Tg and the 5xFAD amyloidosis model indicates that the RNA splicing defect synergizes with the amyloid cascade to remodel the brain transcriptome and proteome, deregulate synaptic proteins, and accelerate cognitive decline. Thus, our results support the contribution of U1 snRNP-mediated splicing dysfunction to AD pathogenesis.

A Case for Thalamic Mechanisms of Schizophrenia: Perspective From Modeling 22q11.2 Deletion Syndrome.

Schizophrenia is a severe, chronic psychiatric disorder that devastates the lives of millions of people worldwide. The disease is characterized by a constellation of symptoms, ranging from cognitive deficits, to social withdrawal, to hallucinations. Despite decades of research, our understanding of the neurobiology of the disease, specifically the neural circuits underlying schizophrenia symptoms, is still in the early stages. Consequently, the development of therapies continues to be stagnant, and overall prognosis is poor. The main obstacle to improving the treatment of schizophrenia is its multicausal, polygenic etiology, which is difficult to model. Clinical observations and the emergence of preclinical models of rare but well-defined genomic lesions that confer substantial risk of schizophrenia (e.g., 22q11.2 microdeletion) have highlighted the role of the thalamus in the disease. Here we review the literature on the molecular, cellular, and circuitry findings in schizophrenia and discuss the leading theories in the field, which point to abnormalities within the thalamus as potential pathogenic mechanisms of schizophrenia. We posit that synaptic dysfunction and oscillatory abnormalities in neural circuits involving projections from and within the thalamus, with a focus on the thalamocortical circuits, may underlie the psychotic (and possibly other) symptoms of schizophrenia.

Hypoxia alters the release and size distribution of extracellular vesicles in pancreatic cancer cells to support their adaptive survival.

Pancreatic tumors are highly desmoplastic and poorly-vascularized, and therefore must develop adaptive mechanisms to sustain their survival under hypoxic condition. Extracellular vesicles (EV) play vital roles in pancreatic tumor pathobiology by facilitating intercellular communication. Here we studied the effect of hypoxia on the release of EVs and examined their role in adaptive survival of pancreatic cancer (PC) cells. Hypoxia promoted the release of EV in PC cell lines, MiaPaCa and AsPC1, wherein former exhibited a far greater induction. Moreover, a time-dependent, measurable and significant increase was recorded for small EV (SEV) in both the cell lines with only minimal induction observed for medium (MEV) and large EVs (LEV). Similarly, noticeable changes in size distribution of SEV were also recorded with a shift toward smaller average size under extreme hypoxia. Thrombospondin (apoptotic bodies marker) was exclusively detected on LEVs, while Arf6 (microvesicles marker) was mostly present on MEV with some expression in LEV as well. However, CD9 and CD63 (exosome markers) were expressed in both SEV and MEVs with a decreased expression recorded under hypoxia. Among all subfractions, SEV was the most bioactive in promoting the survival of hypoxic PC cells and hypoxia-inducible factor-1α stabilization was involved in heightened EV release under hypoxia and for their potency to promote hypoxic cell survival. Altogether, our findings provide a novel mechanism for the adaptive hypoxic survival of PC cells and should serve as the basis for future investigations on broader functional implications of EV.

Exosomal markers (CD63 and CD9) expression and their prognostic significance using immunohistochemistry in patients with pancreatic ductal adenocarcinoma.

BACKGROUND: Exosomes are important mediators of intercellular communications and play pivotal roles in cancer progression, metastasis and chemoresistance. CD63 and CD9 are widely accepted exosomal markers. In patients with pancreatic ductal adenocarcinoma (PDAC), positive correlation between CD9 expression and overall survival (OS) was reported. CD63 expression was conserved in all patients with no reported prognostic significance. This study explored the prognostic significance of CD63 and CD9 expression using immunohistochemistry (IHC) in patients with PDAC of mixed racial background. METHODS: Between 2012 and 2016, 49 patients with PDAC had available tissues for CD63 and CD9 staining using IHC. Two pathologists independently scored the CD63 and CD9 expression. Staining intensity was graded from 1-3 and staining percentage was estimated in 10% increments. Mean Quick-score (Q-score) (Intensity X Percentage of staining) was calculated. RESULTS: The mean Q-score for CD63 and CD9 are higher in primary tumor from the pancreas compared to pancreatic tumor from metastatic sites (185 vs. 102, P=0.0002) and (48 vs. 20, P=0.0418) respectively. We fitted Cox proportion hazard regression models to investigate the impact of the covariates CD63 and CD9 on progression free survival (PFS) and OS. CD63 has significant impact on PFS (P=0.0135) and OS (P=0.003). The higher the CD63 Q-score, the longer the PFS and OS. CD9 doesn't have significant impact on PFS (P=0.5734) or OS (P=0.2682). The mean CD63 and CD9 Q-scores are slightly higher in African American (AA) compared to Caucasians (157 vs. 149, P=0.76) and (45 vs. 29, P=0.43) respectively. CONCLUSIONS: CD63 and CD9 expression is higher in primary tumor from the pancreas compared to pancreatic tumor from metastatic sites. There is correlation between CD63 expression (but not CD9 in this cohort) and PFS and OS. To our knowledge, this is the first study to show prognostic significance of CD63 expression in patients with PDAC using IHC. A trend of higher expression of CD63 and CD9 among AA compared to Caucasians was also noticed.

Ensemble encoding of action speed by striatal fast-spiking interneurons.

Striatal fast-spiking interneurons (FSIs) potently inhibit the output neurons of the striatum and, as such, powerfully modulate action learning. Through electrical synaptic coupling, FSIs are theorized to temporally coordinate their activity. This has important implications for their ability to temporally summate inhibition on downstream striatal projection neurons. While some in vivo single-unit electrophysiological recordings of putative FSIs support coordinated firing, others do not. Moreover, it is unclear as to what aspect of action FSIs encode. To address this, we used in vivo calcium imaging of genetically identified FSIs in freely moving mice and applied machine learning analyses to decipher the relationship between FSI activity and movement. We report that FSIs exhibit ensemble activity that encodes the speed of action sub-components, including ambulation and head movements. These results suggest FSI population dynamics fit within a Hebbian model for ensemble inhibition of striatal output guiding action.

TrkB-dependent disinhibition of the nucleus accumbens is enhanced by ethanol.

The nucleus accumbens is a critical integration center for reward-related circuitry and is comprised primarily of medium spiny projection neurons. The dynamic balance of excitation and inhibition onto medium spiny neurons determines the output of this structure. While nucleus accumbens excitatory synaptic plasticity is well-characterized, inhibitory synaptic plasticity mechanisms and their potential relevance to shaping motivated behaviors is poorly understood. Here we report the discovery of long-term depression of inhibitory synaptic transmission in the mouse nucleus accumbens core. This long-term depression is postsynaptically expressed, tropomyosin kinase B (TrkB) receptor-mediated, and augmented in the presence of ethanol. Our findings support the emerging view that TrkB signaling regulates inhibitory synaptic plasticity and suggest this mechanism in the nucleus accumbens as a target for ethanol modulation of reward.

Activation of the Rostral Intralaminar Thalamus Drives Reinforcement through Striatal Dopamine Release.

Glutamatergic projections of the thalamic rostral intralaminar nuclei of the thalamus (rILN) innervate the dorsal striatum (DS) and are implicated in dopamine (DA)-dependent incubation of drug seeking. However, the mechanism by which rILN signaling modulates reward seeking and striatal DA release is unknown. We find that activation of rILN inputs to the DS drives cholinergic interneuron burst-firing behavior and DA D2 receptor-dependent post-burst pauses in cholinergic interneuron firing. In vivo, optogenetic activation of this pathway drives reinforcement in a DA D1 receptor-dependent manner, and chemogenetic suppression of the rILN reduces dopaminergic nigrostriatal terminal activity as measured by fiber photometry. Altogether, these data provide evidence that the rILN activates striatal cholinergic interneurons to enhance the pursuit of reward through local striatal DA release and introduce an additional level of complexity in our understanding of striatal DA signaling.

Rejuvenation of plasticity in the brain: opening the critical period.

Cortical circuits are particularly sensitive to incoming sensory information during well-defined intervals of postnatal development called 'critical periods'. The critical period for cortical plasticity closes in adults, thus restricting the brain's ability to indiscriminately store new sensory information. For example, children acquire language in an exposure-based manner, whereas learning language in adulthood requires more effort and attention. It has been suggested that pairing sounds with the activation of neuromodulatory circuits involved in attention reopens this critical period. Here, we review two critical period hypotheses related to neuromodulation: cortical disinhibition and thalamic adenosine. We posit that these mechanisms co-regulate the critical period for auditory cortical plasticity. We also discuss ways to reopen this period and rejuvenate cortical plasticity in adults.

An Aplysia-like synaptic switch for rapid protection against ethanol-induced synaptic inhibition in a mammalian habit circuit.

Decades of work in Aplysia californica established the general rule that principles of synaptic plasticity and their molecular mechanisms are evolutionarily conserved from mollusks to mammals. However, an exquisitely sensitive, activity-dependent homosynaptic mechanism that protects against the depression of neurotransmitter release in Aplysia sensory neuron terminals has, to date, not been uncovered in other animals, including mammals. Here, we discover that depression at a mammalian synapse that is implicated in habit formation and habit learning acceleration by ethanol, the fast-spiking interneuron (FSI) to medium spiny principal projection neuron (MSN) synapse of the dorsolateral striatum, is subject to this type of synaptic protection. We show that this protection against synaptic depression is calcium- and PDZ domain interaction-dependent. These findings support activity dependent protection against synaptic depression as an Aplysia-like synaptic switch in mammals that may represent a leveraging point for treating alcohol use disorders.

Understanding functional and social risk characteristics of frail older adults: a cross-sectional survey study.

BACKGROUND: Frailty is a condition of increasing importance, given the aging adult population. With an anticipated shortage of geriatricians, primary care physicians will increasingly need to manage care for frail adults with complex functional risks and social-economic circumstances. METHODS: We used cross-sectional data from 4551 adults ages 65-90 who responded to the 2014/2015 cycle of the Kaiser Permanente Northern California Member Health Survey (MHS), a self-administered survey that covers multiple health and social characteristics, to create a deficits accumulation model frailty index, classify respondents as frail or non-frail, and then compare prevalence of functional health issues including Activities of Daily Living (ADL)/Instrumental Activities of Daily Living (IADL) and social determinants of health (SDOHs) by frailty status. RESULTS: The overall prevalence of frailty was 14.3%, higher for women than men, increased with age, and more common among those with low levels of education and income. Frail older adults were more likely than non-frail to have ≥ 3 chronic diseases (55.9% vs. 10.1%), obesity (32.7% vs. 22.8%), insomnia (36.4% vs. 8.8%), oral health problems (25.1% vs. 4.7%), balance or walking problems (54.2% vs. 4.9%), ≥ 1 fall (56.1% vs. 19.7%), to use ≥ 1 medication known to increase fall risk (56.7% vs. 26.0%), and to need help with ≥2 ADLs (15.8% vs. 0.8%) and ≥ 2 IADLs (38.4% vs. 0.8%). They were more likely to feel financial strain (26.9% vs. 12.6%) and to use less medication than prescribed (7.4% vs. 3.6%), less medical care than needed (8.3% vs 3.7%), and eat less produce (9.5% vs. 3.2%) due to cost. Nearly 20% of frail adults were unpaid caregivers for an adult with frailty, serious illness or disability. CONCLUSIONS: This study examined the prevalence of frailty and identified modifiable and non-modifiable risk factors of health. The frail older adult population is heterogeneous and requires a patient-centered assessment of their circumstances by healthcare providers and caregivers to improve their quality of life, avoid adverse health events, and slow physical and mental decline. The characteristics identified in this study can be proactively used for the assessment of patient health, quality of life, and frailty prevention.

Complex Control of Striatal Neurotransmission by Nicotinic Acetylcholine Receptors via Excitatory Inputs onto Medium Spiny Neurons.

The prevalence of nicotine dependence is higher than that for any other substance abuse disorder; still, the underlying mechanisms are not fully established. To this end, we studied acute effects by nicotine on neurotransmission in the dorsolateral striatum, a key brain region with respect to the formation of habits. Electrophysiological recordings in acutely isolated brain slices from rodent showed that nicotine (10 nm to 10 µm) produced an LTD of evoked field potentials. Current-clamp recordings revealed no significant effect by nicotine on membrane voltage or action potential frequency, indicating that the effect by nicotine is primarily synaptic. Nicotine did not modulate sIPSCs, or the connectivity between fast-spiking interneurons and medium spiny neurons, as assessed by whole-cell recordings combined with optogenetics. However, the frequency of sEPSCs was significantly depressed by nicotine. The effect by nicotine was mimicked by agonists targeting α7- or α4-containing nAChRs and blocked in slices pretreated with a mixture of antagonists targeting these receptor subtypes. Nicotine-induced LTD was furthermore inhibited by dopamine D2 receptor antagonist and occluded by D2 receptor agonist. In addition, modulation of cholinergic neurotransmission suppressed the responding to nicotine, which might reflect upon the postulated role for nAChRs as a presynaptic filter to differentially govern dopamine release depending on neuronal activity. Nicotine-induced suppression of excitatory inputs onto medium spiny neurons may promote nicotine-induced locomotor stimulation and putatively initiate neuroadaptations that could contribute to the transition toward compulsive drug taking.SIGNIFICANCE STATEMENT To decrease smoking, prevalence factors that may contribute to the development of nicotine addiction need to be identified. The data presented here show that nicotine suppresses striatal neurotransmission by selectively reducing the frequency of excitatory inputs to medium spiny neurons (MSNs) while rendering excitability, inhibitory neurotransmission, and fast-spiking interneuron-MSN connectivity unaltered. In addition, we show that the effect displayed by nicotine outlasts the presence of the drug, which could be fundamental for the addictive properties of nicotine. Considering the inhibitory tone displayed by MSNs on dopaminergic cell bodies and local terminals, nicotine-induced long-lasting depression of striatal output could play a role in behavioral transformations associated with nicotine use, and putatively elicit neuroadaptations underlying compulsive drug-seeking habits.

Drp1 Mitochondrial Fission in D1 Neurons Mediates Behavioral and Cellular Plasticity during Early Cocaine Abstinence.

Altered brain energy homeostasis is a key adaptation occurring in the cocaine-addicted brain, but the effect of cocaine on the fundamental source of energy, mitochondria, is unknown. We demonstrate an increase of dynamin-related protein-1 (Drp1), the mitochondrial fission mediator, in nucleus accumbens (NAc) after repeated cocaine exposure and in cocaine-dependent individuals. Mdivi-1, a demonstrated fission inhibitor, blunts cocaine seeking and locomotor sensitization, while blocking c-Fos induction and excitatory input onto dopamine receptor-1 (D1) containing NAc medium spiny neurons (MSNs). Drp1 and fission promoting Drp1 are increased in D1-MSNs, consistent with increased smaller mitochondria in D1-MSN dendrites after repeated cocaine. Knockdown of Drp1 in D1-MSNs blocks drug seeking after cocaine self-administration, while enhancing the fission promoting Drp1 enhances seeking after long-term abstinence from cocaine. We demonstrate a role for altered mitochondrial fission in the NAc, during early cocaine abstinence, suggesting potential therapeutic treatment of disrupting mitochondrial fission in cocaine addiction.

Incidence, duration and cost of futile treatment in end-of-life hospital admissions to three Australian public-sector tertiary hospitals: a retrospective multicentre cohort study.

OBJECTIVES: To estimate the incidence, duration and cost of futile treatment for end-of-life hospital admissions. DESIGN: Retrospective multicentre cohort study involving a clinical audit of hospital admissions. SETTING: Three Australian public-sector tertiary hospitals. PARTICIPANTS: Adult patients who died while admitted to one of the study hospitals over a 6-month period in 2012. MAIN OUTCOME MEASURES: Incidences of futile treatment among end-of-life admissions; length of stay in both ward and intensive care settings for the duration that patients received futile treatments; health system costs associated with futile treatments; monetary valuation of bed days associated with futile treatment. RESULTS: The incidence rate of futile treatment in end-of-life admissions was 12.1% across the three study hospitals (range 6.0%-19.6%). For admissions involving futile treatment, the mean length of stay following the onset of futile treatment was 15 days, with 5.25 of these days in the intensive care unit. The cost associated with futile bed days was estimated to be $AA12.4 million for the three study hospitals using health system costs, and $A988 000 when using a decision maker's willingness to pay for bed days. This was extrapolated to an annual national health system cost of $A153.1 million and a decision maker's willingness to pay of $A12.3 million. CONCLUSIONS: The incidence rate and cost of futile treatment in end-of-life admissions varied between hospitals. The overall impact was substantial in terms of both the bed days and cost incurred. An increased awareness of these economic costs may generate support for interventions designed to reduce futile treatments. We did not include emotional hardship or pain and suffering, which represent additional costs.

Exosomal Markers (CD63 and CD9) Expression Pattern Using Immunohistochemistry in Resected Malignant and Nonmalignant Pancreatic Specimens.

OBJECTIVES: Exosomes are important mediators in intercellular communications and play a role in cancer progression and metastasis. Exosomal membranes are enriched in endosome-specific tetraspanins (CD9 and CD63). Here, we explored the expression of CD63 and CD9 utilizing immunohistochemistry in malignant and nonmalignant cells in 29 resected pancreatic specimens (RPSs) of mixed racial background. METHODS: The pathologic tissues (PTs) and adjacent normal tissues (ANTs) in each RPS were stained for CD63 and CD9. Two pathologists independently scored the expression of CD63 and CD9. Staining intensity was graded from 1 to 3. Staining percentage was estimated in 10% increments. An average Quick score (Q score) (intensity × percentage of staining) was calculated. Unpaired t test was used for statistical analysis. RESULTS: The mean multiplicative Q score for CD63 and CD9 expression is higher in PTs (209 and 72) compared with ANTs (154 and 24) (P = 0.0041, P = 0.0018), respectively. The Mean Q score for CD63 and CD9 expression is higher in the malignant PTs (231 and 85) compared with ANTs (129 and 25) (P < 0.0001 and P < 0.0124). CONCLUSIONS: Exosomal markers (CD63 and CD9) expression assessment using immunohistochemistry is feasible in RPS. The expression of CD63 and CD9 is higher in PTs and malignant PTs compared with their ANTs.