Preclinical Research Articles

Feasibility of a novel augmented reality overlay for cervical screw placement in phantom spine models.

Feasibility study. A phantom model was used to evaluate the accuracy of a novel augmented reality (AR) system for cervical screw placement. The use of navigation systems is becoming increasingly common in spine procedures. However, numerous factors limit the feasibility of regular and widespread use of navigation tools during spine surgery. AR is a new technology that has already demonstrated utility as a navigation tool during spine surgery. However, advancements in AR technology are needed to increase its adoption by the medical community. AR technology that uses a fiducial-less registration system was tested in a preclinical cervical spine phantom model study for accuracy during spinal screw placement. A three-dimensional reconstruction of the spine along with trajectory lines was superimposed onto the phantom model using an AR headset. Participants used the AR system to guide screw placement, and post-instrumentation scans were compared for accuracy assessment. Twelve cervical screws were placed under AR guidance. All screws were placed in an acceptable anatomic position. The average distance error for the insertion point was 2.73±0.55 mm, whereas that for the endpoint was 2.71±0.69 mm. The average trajectory angle error for all insertions was 2.69°±0.59°. This feasibility study describes a novel registration approach that superimposes spinal anatomy and trajectories onto the surgeon's real-world view of the spine. These results demonstrate reasonable accuracy in the preclinical model. The results of this study demonstrate that this technology can assist with accurate screw placement. Further investigation using cadaveric and clinical models is warranted.

Relapse after intermittent access to cocaine: Discriminative cues more effectively trigger drug seeking than do conditioned cues.

When people with drug addiction encounter cues associated with drug use, this can trigger cravings and relapse. These cues can include conditioned stimuli (CSs) signaling drug delivery and discriminative stimuli (DSs) signaling drug availability. Compared to CS effects, DS effects are less explored in preclinical studies on cue-induced relapse. We compared CS and DS effects on reward seeking following abstinence from intermittent-access cocaine (or sucrose) self-administration. During 15-20 intermittent-access sessions, rats self-administered i.v. cocaine or sucrose pellets paired with a light-tone CS. Cocaine/sucrose was available for 5-min (signalled by a light; DS+) and unavailable for 25min (signalled by different lighting conditions; DS-), and this cycled for 4h/session. Following abstinence, we measured cocaine/sucrose seeking under extinction triggered by CS and DS presentation, and instrumental responding reinforced by these cues. Across intermittent-access sessions, rats increased lever pressing for cocaine or sucrose during DS+ periods and decreased responding during DS- periods. On days 2 and 21 of abstinence, only presentation of the DS+ or DS+ and CS combined elicited increased cocaine/sucrose-seeking behaviour (i.e., increased active lever presses). Presenting the DS- alongside the DS+ suppressed the increased cocaine-seeking behaviour otherwise produced by the DS+ . Finally, on day 21 of abstinence, rats showed equivalent levels of lever pressing reinforced by the DS+ , CS and by the DS+ and CS combined, suggesting comparable conditioned reinforcing value. After intermittent self-administration, cocaine-associated DSs and CSs acquire similar conditioned reinforcing properties, but DSs more effectively trigger increases in drug seeking.

Personalized Nanovaccines Enhance Lymph Node Accumulation and Reprogram the Tumor Microenvironment for Improved Photodynamic Immunotherapy.

Tumor immunotherapy has emerged as an efficacious therapeutic approach that mobilizes the patient's immune system to achieve durable tumor suppression. Here, we design a photodynamic therapy-motivated nanovaccine (Dex-HDL/ALA-Fe3O4) co-delivering 5-aminolevulinic acid and Fe3O4 nanozyme that demonstrate a long-term durable immunotherapy strategy. After vaccination, the nanovaccine exhibits obvious tumor site accumulation, lymph node homing, and specific and memory antitumor immunity evocation. Upon laser irradiation, Dex-HDL/ALA-Fe3O4 effectively generates reactive oxygen species at the tumor site not only to induce the immunogenic cell death-cascade but also to trigger the on-demand release of full types of tumor antigens. Intriguingly, Fe3O4 nanozyme-catalyzed hydrogen peroxide generated oxygen for alleviating tumor hypoxia and modifying the inhibitory tumor microenvironment, thereby exhibiting remarkable potential as a sensitizer. The intravenous administration of nanovaccines in diverse preclinical cancer models has demonstrated remarkable tumor regression and inhibition of postoperative tumor recurrence and metastasis, thereby enabling personalized treatment strategies against highly heterogeneous tumors.

Exogenous Ketone Esters as a Potential Therapeutic for Treatment of Sarcopenic Obesity.

Identifying effective treatment(s) for sarcopenia and sarcopenic obesity are of paramount importance as the global population advances in age and obesity continues to be a worldwide concern. Evidence has shown that a ketogenic diet can be beneficial for preservation of muscle quality and function in older adults, but long-term adherence is low due in part to the high-fat (> 80%), very low carbohydrate (< 5%) composition of the diet. When provided in adequate amounts, exogenous ketone esters can increase circulating ketones to concentrations that exceed those observed during prolonged fasting or starvation without significant alterations in the diet. Ketone esters first emerged in the mid-1990s and their use in pre-clinical and clinical research has escalated within the past 10-15 years. We present findings from a narrative review of the existing literature for a proposed hypothesis on the effects of exogenous ketones as a therapeutic for preservation of skeletal muscle and function within the context of sarcopenic obesity and future directions for exploration. Much of the reviewed literature herein examines the mechanisms of the ketone diester (R, S-1,3-butanediol diacetoacetate) on skeletal muscle mass, muscle protein synthesis, and epigenetic regulation in murine models. Additional studies are needed to further examine the key regulatory factors producing these effects in skeletal muscle, examine convergent and divergent effects among different ketone ester formulations, and establish optimal frequency and dosing regimens to translate these findings into humans.

Biomimetic chemical reactions with natural products using metalloporphyrins and salen complexes as catalysts: a brief review.

The cytochrome P450 is a superfamily of hemoproteins mainly present in the liver and are versatile biocatalysts. They participate in the primary metabolism and biosynthesis of various secondary metabolites. Chemical catalysts are utilized to replicate the activities of enzymes. Metalloporphyrins and Salen complexes can contribute to the products' characterization and elucidate biotransformation processes, which are investigated during pre-clinical trials. These catalysts also help discover biologically active compounds and get better yields of products of industrial interest. This review aims to investigate which natural product classes are being investigated by biomimetic chemical models and the functionalities applied in the use of these catalysts. A limited number of studies were observed, with terpenes and alkaloids being the most investigated natural product classes. The research also revealed that Metalloporphyrins are still the most popular in the studies, and the identity and yield of the products obtained depend on the reaction system conditions.

Histological evaluation of osseointegration between conventional and novel bone-level tapered implants in healed bone-A preclinical study.

To histologically compare osseointegration and crestal bone healing between newly introduced tapered, self-cutting bone-level test implants and tapered bone-level control implants in sites with fully healed sites. Sixty-six implants (33 test, 33 control) were placed 1 mm subcrestally in a minipig model and underwent qualitative histologic and quantitative histometric analyses after 3, 6 and 12 weeks of submerged healing. The primary and secondary outcomes were the bone-to-implant contact (BIC) and first bone-to-implant contact (fBIC). Outcomes between the test and control implants were statistically compared. The BIC values of the test implants were comparable and non-inferior over the time points studied, except for the 12 weeks time point which showed statistically significantly higher BIC values of the test (88.07 ± 5.35%) compared to the control implants (80.88 ± 7.51%) (p = .010). Similarly comparable and non-inferior were the fBIC values, except for the 6-week outcome, which showed statistically higher values for the test (-546.5 ± 450.80 μm) compared to the control implants (-75.7 ± 100.59 μm). fBIC results for the test implants were qualitatively more stable and consistent between test time points. Novel self-cutting bone-level test implants demonstrated superior osseointegration and similar bone levels compared to conventional bone-level implants after a healing period of 12 weeks in healed ridges.

VEGF-R2/CAV-1 Interaction Induced by Resveratrol/Eicosapentaenoic Acid/Docosahexaenoic Acid-Enriched Formulation through Functional Detergent-Resistant Membranes Is Associated with Decreased VEGF-A Release in ARPE-19 Cells.

Omega-3 fatty acids (O3FAs) and resveratrol (RSV) are known to be beneficial for certain eye diseases, such as age-related macular degeneration (AMD). Neovascular AMD is characterized by abnormal blood vessel formation due to the excessive synthesis of vascular endothelial growth factor (VEGF) by retinal pigment epithelium (RPE) cells. The study investigates whether a formulation based on eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), and RSV is capable of counteracting VEGF-A secretion, and elucidates the molecular mechanism. The study finds, using ELISA, that O3FAs/RSV reduces VEGF-A secretion in human RPE cells. This phenomenon is related to the increased interaction between VEGF-receptor 2 (VEGF-R2) and caveolin-1 (CAV-1), a protein of detergent-resistant membranes (DRMs), as determined by co-immunoprecipitation and proximity ligation assay. Using microscale thermophoresis, the study confirms that O3FAs/RSV causes a high-affinity interaction. Isolation and analysis of DRMs reveal that this interaction is concomitant with VEGF-R2 relocalization in DRMs. The depletion of DRMs by a cholesterol-chelating agent blocks the VEGF-R2/CAV-1 interaction and EPA/DHA/RSV-mediated impairment of VEGF production. This specific interaction can provide a new strategy for countering VEGF-A production in human RPE cells and, consequently, reducing neovascularization in AMD. Further preclinical studies involving O3FAs and polyphenols are warranted.

Insights into Nimbolide molecular crosstalk and its anticancer properties.

Nimbolide, one of the main ingredients constituent of Azadirachta indica (neem) leaf extract, has garnered attention for its potential as an anticancer agent. Its efficacy against various cancers and chemopreventive action has been demonstrated through numerous in vivo and in vitro studies. This updated review aims to comprehensively explore the chemopreventive and anticancer properties of nimbolide, emphasizing its molecular mechanisms of action and potential therapeutic applications in oncology. The review synthesizes evidence from various studies that examine nimbolide's roles in apoptosis induction, anti-proliferation, cell death, metastasis inhibition, angiogenesis suppression, and modulation of carcinogen-metabolizing enzymes. Nimbolide exhibits multifaceted anticancer activities, including the modulation of multiple cell signaling pathways related to inflammation, invasion, survival, growth, metastasis, and angiogenesis. However, its pharmacological development is still in the early stages, mainly due to limited pharmacokinetic and comprehensive long-term toxicological studies. Nimbolide shows promising anticancer and chemopreventive properties, but there is need for systematic preclinical pharmacokinetic and toxicological research. Such studies are essential for establishing safe dosage ranges for first-in-human clinical trials and further advancing nimbolide's development as a therapeutic agent against various cancers. The review highlights the potential of nimbolide in cancer treatment and underscores the importance of rigorous preclinical evaluation to realize its full therapeutic potential.

A prognostic neural epigenetic signature in high-grade glioma.

Neural-tumor interactions drive glioma growth as evidenced in preclinical models, but clinical validation is limited. We present an epigenetically defined neural signature of glioblastoma that independently predicts patients' survival. We use reference signatures of neural cells to deconvolve tumor DNA and classify samples into low- or high-neural tumors. High-neural glioblastomas exhibit hypomethylated CpG sites and upregulation of genes associated with synaptic integration. Single-cell transcriptomic analysis reveals a high abundance of malignant stemcell-like cells in high-neural glioblastoma, primarily of the neural lineage. These cells are further classified as neural-progenitor-cell-like, astrocyte-like and oligodendrocyte-progenitor-like, alongside oligodendrocytes and excitatory neurons. In line with these findings, high-neural glioblastoma cells engender neuron-to-glioma synapse formation in vitro and in vivo and show an unfavorable survival after xenografting. In patients, a high-neural signature is associated with decreased overall and progression-free survival. High-neural tumors also exhibit increased functional connectivity in magnetencephalography and resting-state magnet resonance imaging and can be detected via DNA analytes and brain-derived neurotrophic factor in patients' plasma. The prognostic importance of the neural signature was further validated in patients diagnosed with diffuse midline glioma. Our study presents an epigenetically defined malignant neural signature in high-grade gliomas that is prognostically relevant. High-neural gliomas likely require a maximized surgical resection approach for improved outcomes.

IDH inhibition in gliomas: from preclinical models to clinical trials.

Gliomas are the most common malignant primary brain tumours in adults and cannot usually be cured with standard cancer treatments. Gliomas show intratumoural and intertumoural heterogeneity at the histological and molecular levels, and they frequently contain mutations in the isocitrate dehydrogenase 1 (IDH1) or IDH2 gene. IDH-mutant adult-type diffuse gliomas are subdivided into grade 2, 3 or 4 IDH-mutant astrocytomas and grade 2 or 3 IDH-mutant, 1p19q-codeleted oligodendrogliomas. The product of the mutated IDH genes, D-2-hydroxyglutarate (D-2-HG), induces global DNA hypermethylation and interferes with immunity, leading to stimulation of tumour growth. Selective inhibitors of mutant IDH, such as ivosidenib and vorasidenib, have been shown to reduce D-2-HG levels and induce cellular differentiation in preclinical models and to induce MRI-detectable responses in early clinical trials. The phase III INDIGO trial has demonstrated superiority of vorasidenib, a brain-penetrant pan-mutant IDH inhibitor, over placebo in people with non-enhancing grade 2 IDH-mutant gliomas following surgery. In this Review, we describe the pathway of development of IDH inhibitors in IDH-mutant low-grade gliomas from preclinical models to clinical trials. We discuss the practice-changing implications of the INDIGO trial and consider new avenues of investigation in the field of IDH-mutant gliomas.

Nutrition and chronic musculoskeletal pain: A narrative review and directions for temporomandibular disorder research and management.

Recent evidence suggests neuro-immune mechanisms may link dietary patterns to chronic painful conditions (CPC). In the research field of oro-facial pain (OFP), studies focuses primarily on dietary mechanical limitations due to pain and dysfunction. This narrative review aimed to overview the role of nutrition on CPC, with emphasis on temporomandibular disorder (TMD), enlightening OFP researcher on dietary assessment possibilities and providing directions for studies in the field of OFP and nutrition. A PubMed database search was performed using the MeSH and non-MeSH descriptors: "temporomandibular joint disorder"; "orofacial pain"; "musculoskeletal pain"; "chronic pain disorders"; "nutrition"; "diet"; "dietary therapy"; "dietary intake" and "inflammation". No time restrictions were applied. Literature reviews, systematic reviews, meta-analyses and clinical and pre-clinical trials were included. Exogenous oxidants from unhealthy dietary patterns may contribute to peripheral and central pro-inflammatory immune signalling leading to peripheral and central sensitization. Furthermore, diets rich in bioactive compounds are suggested to contribute to pain management of CPC. High dietary intake of ultra-processed foods impacts the quality of the diet and shows adverse health outcomes. In this context, the role of nutrition on TMD remains overlooked. Considering diet may influence CPC, allied with the scarcity of studies evaluating the role of nutrition on TMD, well-designed clinical trials based on dietary assessments and measurements capable of evaluating food quality, UPF consumption and nutrient adequacy-added to serum nutrient levels evaluation-are suggested.

Radiation enhancement using focussed ultrasound-stimulated microbubbles for breast cancer: A Phase 1 clinical trial.

Preclinical studies have demonstrated that tumour cell death can be enhanced 10- to 40-fold when radiotherapy is combined with focussed ultrasound-stimulated microbubble (FUS-MB) treatment. The acoustic exposure of microbubbles (intravascular gas microspheres) within the target volume causes bubble cavitation, which induces perturbation of tumour vasculature and activates endothelial cell apoptotic pathways responsible for the ablative effect of stereotactic body radiotherapy. Subsequent irradiation of a microbubble-sensitised tumour causes rapid increased tumour death. The study here presents the mature safety and efficacy outcomes of magnetic resonance (MR)-guided FUS-MB (MRgFUS-MB) treatment, a radioenhancement therapy for breast cancer. This prospective, single-center, single-arm Phase 1 clinical trial included patients with stages I-IV breast cancer with in situ tumours for whom breast or chest wall radiotherapy was deemed adequate by a multidisciplinary team (clinicaltrials.gov identifier: NCT04431674). Patients were excluded if they had contraindications for contrast-enhanced MR or microbubble administration. Patients underwent 2 to 3 MRgFUS-MB treatments throughout radiotherapy. An MR-coupled focussed ultrasound device operating at 800 kHz and 570 kPa peak negative pressure was used to sonicate intravenously administrated microbubbles within the MR-guided target volume. The primary outcome was acute toxicity per Common Terminology Criteria for Adverse Events (CTCAE) v5.0. Secondary outcomes were tumour response at 3 months and local control (LC). A total of 21 female patients presenting with 23 primary breast tumours were enrolled and allocated to intervention between August/2020 and November/2022. Three patients subsequently withdrew consent and, therefore, 18 patients with 20 tumours were included in the safety and LC analyses. Two patients died due to progressive metastatic disease before 3 months following treatment completion and were excluded from the tumour response analysis. The prescribed radiation doses were 20 Gy/5 fractions (40%, n = 8/20), 30 to 35 Gy/5 fractions (35%, n = 7/20), 30 to 40 Gy/10 fractions (15%, n = 3/20), and 66 Gy/33 fractions (10%, n = 2/20). The median follow-up was 9 months (range, 0.3 to 29). Radiation dermatitis was the most common acute toxicity (Grade 1 in 16/20, Grade 2 in 1/20, and Grade 3 in 2/20). One patient developed grade 1 allergic reaction possibly related to microbubbles administration. At 3 months, 18 tumours were evaluated for response: 9 exhibited complete response (50%, n = 9/18), 6 partial response (33%, n = 6/18), 2 stable disease (11%, n = 2/18), and 1 progressive disease (6%, n = 1/18). Further follow-up of responses indicated that the 6-, 12-, and 24-month LC rates were 94% (95% confidence interval [CI] [84%, 100%]), 88% (95% CI [75%, 100%]), and 76% (95% CI [54%, 100%]), respectively. The study's limitations include variable tumour sizes and dose fractionation regimens and the anticipated small sample size typical for a Phase 1 clinical trial. MRgFUS-MB is an innovative radioenhancement therapy associated with a safe profile, potentially promising responses, and durable LC. These results warrant validation in Phase 2 clinical trials. clinicaltrials.gov, identifier NCT04431674.

Microbial- and host immune cell-derived extracellular vesicles in the pathogenesis and therapy of periodontitis: A narrative review.

Periodontitis is a chronic inflammatory disease caused by dysbiotic biofilms and destructive host immune responses. Extracellular vesicles (EVs) are circulating nanoparticles released by microbes and host cells involved in cell-to-cell communication, found in body biofluids, such as saliva and gingival crevicular fluid (GCF). EVs are mainly involved in cell-to-cell communication,and may hold promise for diagnostic and therapeutic purposes. Periodontal research has examined the potential involvement of bacterial- and host-cell-derived EVs in disease pathogenesis, diagnosis, and therapy, but data remains scarce on immune cell- or microbial-derived EVs. In this narrative review, we first provide an overview of the role of microbial and host-derived EVs on disease pathogenesis. Recent studies reveal that Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans-derived outer membrane vesicles (OMVs) can activate inflammatory cytokine release in host cells, while M1 macrophage EVs may contribute to bone loss. Additionally, we summarised current invitro and pre-clinical research on the utilisation of immune cell and microbial-derived EVs as potential therapeutic tools in the context of periodontal treatment. Studies indicate that EVs from M2 macrophages and dendritic cells promote bone regeneration in animal models. While bacterial EVs remain underexplored for periodontal therapy, preliminary research suggests that P. gingivalis OMVs hold promise as vaccine candidates. Finally, we acknowledge the current limitations present in the field of translating immune cell derived EVs and microbial derived EVs in periodontology. It is concluded that microbial and host immune cell-derived EVs have a role in periodontitis pathogenesis and hence may be useful for studying disease pathophysiology, and as diagnostic and treatment monitoring biomarkers.

Therapeutic Promises of Ferulic Acid and Its Derivatives on Hepatic damage Related with Oxidative Stress and Inflammation: A Review with Mechanisms.

Ferulic acid (FA) is a naturally occurring phenolic compound commonly found in the plant Ferula communis. This study aims to investigate the hepatoprotective effect of FA and its derivatives (methyl ferulic acid and trans-ferulic acid) against oxidative stress and inflammation-related hepatotoxicity due to toxicants based on the results of different non-clinical and preclinical tests. For this, data was collected from different reliable electronic databases such as PubMed, Google Scholar, and ScienceDirect, etc. The results of this investigation demonstrated that FA and its derivatives have potent hepatoprotective effects against oxidative stress and inflammation-related damage. The findings also revealed that these protective effects are due to the antioxidant and anti-inflammatory effects of the chemical compound. FA and its analogues significantly inhibit free radical generation and hinder the effects of proinflammatory markers and inflammatory enzymes, resulting in diminished cytotoxic and apoptotic hepatocyte death. The compounds also prevent intracellular lipid accumulation and provide protective effects.

The feasibility of acupuncture on post-spinal cord injury treatment

Spinal cord injury (SCI) simultaneously causes multiple and interrelated pathophysiological disorders throughout the nervous system, hindering the development of effective treatment strategies. Mechanistically, SCI triggers excitatory signaling activation, downregulation of the inhibitory system, neuronal death, followed by the development of new synaptic circuits and reorganization, leading to neurological dysfunctions and mental disorders. Therefore, a simultaneous treatment strategy, known as overlapping treatment, is needed. Over decades, both preclinical and clinical studies have established that acupuncture treatment offers neuroprotection, pain attenuation, improvement of functional recovery, and promotes reward behaviors, suggesting potential roles of acupuncture in post-SCI treatment. Recently, the importance of overlapping treatment has been recognized in developing effective treatments for post-SCI pathophysiology. However, there has been no systematic study investigating the role of acupuncture in various SCI pathophysiology. In this review, we briefly address the mechanisms of post-SCI pathophysiology and discuss the potential therapeutic effects of acupuncture, suggesting its feasibility as a treatment for post-SCI pathophysiology.

Spatial genomic, biochemical and cellular mechanisms underlying meningioma heterogeneity and evolution.

Intratumor heterogeneity underlies cancer evolution and treatment resistance, but targetable mechanisms driving intratumor heterogeneity are poorly understood. Meningiomas are the most common primary intracranial tumors and are resistant to all medical therapies, and high-grade meningiomas have significant intratumor heterogeneity. Here we use spatial approaches to identify genomic, biochemical and cellular mechanisms linking intratumor heterogeneity to the molecular, temporal and spatial evolution of high-grade meningiomas. We show that divergent intratumor gene and protein expression programs distinguish high-grade meningiomas that are otherwise grouped together by current classification systems. Analyses of matched pairs of primary and recurrent meningiomas reveal spatial expansion of subclonal copy number variants associated with treatment resistance. Multiplexed sequential immunofluorescence and deconvolution of meningioma spatial transcriptomes using cell types from single-cell RNA sequencing show decreased immune infiltration, decreased MAPK signaling, increased PI3K-AKT signaling and increased cell proliferation, which are associated with meningioma recurrence. To translate these findings to preclinical models, we use CRISPR interference and lineage tracing approaches to identify combination therapies that target intratumor heterogeneity in meningioma cell co-cultures.

Ameliorative Potential of Bone Marrow-Derived Mesenchymal Stem Cells Versus Prednisolone in a Rat Model of Lung Fibrosis: A Histological, Immunohistochemical, and Biochemical Study.

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease of unknown origin with limited treatment options and poor prognosis. The encouraging findings from preclinical investigations utilizing mesenchymal stem cells (MSCs) indicated that they could serve as a promising therapeutic alternative for managing chronic lung conditions, such as IPF. The objective of this study was to compare the efficiency of bone marrow-derived MSCs (BM-MSCs) versus prednisolone, the standard anti-inflammatory medication, in rats with bleomycin (BLM)-induced lung fibrosis. Four groups were created: a control group, a BLM group, a prednisolone-treated group, and a BM-MSCs-treated group. To induce lung fibrosis, 5 mg/kg of BLM was administered intratracheally. BLM significantly increased serum levels of pro-inflammatory cytokines and oxidative stress markers. The disturbed lung structure was also revealed by light and transmission electron microscopic studies. Upregulation in the immune expression of alpha-smooth muscle actin, transforming growth factor beta-1, and Bax was demonstrated. Interestingly, all findings significantly regressed on treatment with prednisolone and BM-MSCs. However, treatment with BM-MSCs showed better results than with prednisolone. In conclusion, BM-MSCs could be a promising approach for managing lung fibrosis.

Distinctive neurophysiological signatures of analgesia after inflammatory pain in the ACC of freely moving mice.

Preclinical assessments of pain have often relied upon behavioral measurements and anesthetized neurophysiological recordings. Current technologies enabling large scale neural recordings, however, have the potential to unveil quantifiable pain signals in conscious animals for preclinical studies. Although pain processing is distributed across many brain regions, the anterior cingulate cortex (ACC) is of particular interest in isolating these signals given its suggested role in the affective ('unpleasant') component of pain. Here, we explored the utility of the ACC towards preclinical pain research using head-mounted miniaturized microscopes to record calcium transients in freely moving male mice expressing GCaMP6f under the Thy1 promoter. We verified the expression of GCaMP6f in excitatory neurons and found no intrinsic behavioral differences in this model. Using a multimodal stimulation paradigm across naive, pain, and analgesic conditions, we found that while ACC population activity roughly scaled with stimulus intensity, single cell representations were highly flexible. We found only low magnitude increases in population activity after CFA, and insufficient evidence for the existence of a robust nociceptive ensemble in the ACC. However, we found a temporal sharpening of response durations and generalized increases in pairwise neural correlations in the presence of the mechanistically distinct analgesics gabapentin or ibuprofen after (but not before) CFA induced inflammatory pain. This increase was not explainable by changes in locomotion alone. Taken together, these results highlight challenges in isolating distinct pain signals amongst flexible representations in the ACC but suggest a neurophysiological hallmark of analgesia after pain that generalizes to at least two analgesics.Significance Statement Our study measured neural activity in the anterior cingulate cortex (ACC) of transgenic mice to improve measures of pain and analgesia in preclinical models. We found that although ACC population activity scaled with stimulus intensity and could be decoded, single cell representations of sensory stimuli were flexible. Low magnitude increases in ACC population activity were observed after pain, but subpopulations with specific activity changes driven by pain/analgesia were difficult to disambiguate from intrinsic variability. Interestingly, responses were temporally sharpened and exhibited increased cell to cell correlations in the presence of two distinct analgesics after CFA but not before. These distinct neural signatures of analgesia occurring only after pain may broaden our understanding of central mechanisms of pain and analgesia.

Novel mechanisms of action of emerging therapies of hereditary thrombotic thrombocytopenic purpura.

Hereditary thrombotic thrombocytopenic purpura (hTTP) is caused by deficiency of plasma ADAMTS13 activity, resulting from ADAMTS13 mutations. ADAMTS13 cleaves ultra large von Willebrand factor (VWF), thus reducing its multimer sizes. Hereditary deficiency of plasma ADAMTS13 activity leads to the formation of excessive platelet-VWF aggregates in small arterioles and capillaries, resulting in hTTP. PubMed search from 1956-2024 using thrombotic thrombocytopenic purpura and therapy identified 3,675 articles. Only the articles relevant to the topic were selected for discussion, which focuses on pathophysiology, clinical presentations, and mechanisms of action of emerging therapeutics for hTTP. Current therapies include infusion of plasma, or coagulation factor VIII, or recombinant ADAMTS13. Emerging therapies include anti-VWF A1 aptamers or nanobody and gene therapies with adeno-associated viral vector or self-inactivated lentiviral vector or a Sleeping Beauty transposon system for a long-term expression of a functional ADAMTS13 enzyme. Frequent plasma infusion remains to be the standard of care in most parts of the world, while recombinant ADAMTS13 has become the treatment of choice for hTTP in some of the Western countries. The success of gene therapies in preclinical models may hold a promise for future development of these novel approaches for a cure of hTTP.

Brief apnea with hypoventilation reduces seizure duration and shifts seizure location for several hours in a model of severe traumatic brain injury.

Seizures can be difficult to control in infants and toddlers. Seizures with periods of apnea and hypoventilation are common following severe traumatic brain injury (TBI). We previously observed that brief apnea with hypoventilation (A&H) in our severe TBI model acutely interrupted seizures. The current study is designed to determine the effect of A&H on subsequent seizures and whether A&H has potential therapeutic implications. Piglets (1 week or 1 month old) received multifactorial injuries: cortical impact, mass effect, subdural hematoma, subarachnoid hemorrhage, and seizures induced with kainic acid. A&H (1 min apnea, 10 min hypoventilation) was induced either before or after seizure induction, or control piglets received subdural/subarachnoid hematoma and seizure without A&H. In an intensive care unit, piglets were sedated, intubated, and mechanically ventilated, and epidural electroencephalogram was recorded for an average of 18 h after seizure induction. In our severe TBI model, A&H after seizure reduced ipsilateral seizure burden by 80% compared to the same injuries without A&H. In the A&H before seizure induction group, more piglets had exclusively contralateral seizures, although most piglets in all groups had seizures that shifted location throughout the several hours of seizure. After 8-10 h, seizures transitioned to interictal epileptiform discharges regardless of A&H or timing of A&H. Even brief A&H may alter traumatic seizures. In our preclinical model, we will address the possibility of hypercapnia with normoxia, with controlled intracranial pressure, as a therapeutic option for children with status epilepticus after hemorrhagic TBI.