Search

Key Clinical Message Acute Coxiella burnetti (Q fever) infection is known to activate the autoimmune inflammatory response. We report a rare case of glomerulonephritis associated with the Coxiella infection. An elderly male first presented with recurrent fevers of unknown origin and was subsequently diagnosed with Q fever infection and treated with doxycycline. He represented 2 months later with an acute kidney injury and active urinary sediment. Renal biopsy demonstrated a mesangiopathic pattern with staining for both C1q and IgM, raising possibilities of infection‐related glomerulonephritis, C1q nephropathy and lupus nephritis. This case demonstrates glomerulonephritis as a complication of Q fever infection and the differential diagnosis and workup up that needs to be considered in cases of chronic Q fever.

Abstract Organophosphates (OPs) are a class of neurotoxic acetylcholinesterase inhibitors including widely used pesticides as well as nerve agents such as VX and VR. Current treatment of these toxins relies on reactivating acetylcholinesterase, which remains ineffective. Enzymatic scavengers are of interest for their ability to degrade OPs systemically before they reach their target. Here we describe a library of computationally designed variants of phosphotriesterase (PTE), an enzyme that is known to break down OPs. The mutations G208D, F104A, K77A, A80V, H254G, and I274N broadly improve catalytic efficiency of VX and VR hydrolysis without impacting the structure of the enzyme. The mutation I106 A improves catalysis of VR and L271E abolishes activity, likely due to disruptions of PTE's structure. This study elucidates the importance of these residues and contributes to the design of enzymatic OP scavengers with improved efficiency.

Abstract In brown adipose tissue (BAT), short-term cold exposure induces the activating transcription factor 4 (ATF4), and its downstream target fibroblast growth factor 21 (FGF21). Induction of ATF4 in BAT in response to mitochondrial stress is required for thermoregulation, partially by increasing FGF21 expression. In the present study, we tested the hypothesis that Atf4 and Fgf21 induction in BAT are both required for BAT thermogenesis under physiological stress by generating mice selectively lacking either Atf4 (ATF4 BKO) or Fgf21 (FGF21 BKO) in UCP1-expressing adipocytes. After 3 days of cold exposure, core body temperature was significantly reduced in ad-libitum-fed ATF4 BKO mice, which correlated with Fgf21 downregulation in brown and beige adipocytes, and impaired browning of white adipose tissue. Conversely, despite having reduced browning, FGF21 BKO mice had preserved core body temperature after cold exposure. Mechanistically, ATF4, but not FGF21, regulates amino acid import and metabolism in response to cold, likely contributing to BAT thermogenic capacity under ad libitum-fed conditions. Importantly, under fasting conditions, both ATF4 and FGF21 were required for thermogenesis in cold-exposed mice. Thus, ATF4 regulates BAT thermogenesis under fed conditions likely in a FGF21-independent manner, in part via increased amino acid uptake and metabolism.

[This corrects the article DOI: 10.1371/journal.pone.0266608.].

Abstract Background Prostate cancer is the most common cancer among men in the United States. Treatment guidelines recommend active surveillance for low‐risk prostate cancer, which involves monitoring for progression, to avoid or delay definitive treatments and their side effects. Despite increased uptake, adherence to surveillance remains a challenge. Methods We conducted semi‐structured, qualitative, virtual interviews based on the Theoretical Domains Framework (TDF), with men (15) who were or had been on active surveillance for their low‐risk prostate cancer in 2020. Interviews were transcribed and coded under TDF's behavioral theory‐based domains. We analyzed domains related to adherence to surveillance using constructivist grounded theory to identify themes influencing decision processes in adherence. Results The TDF domains of emotion, beliefs about consequences, environmental context and resources, and social influences were most relevant to surveillance adherence‐. From these four TDF domains, three themes emerged as underlying decision processes: trust in surveillance as treatment, quality of life, and experiences of self and others. Positive perceptions of these three themes supported adherence while negative perceptions contributed to non‐adherence (i.e., not receiving follow‐up or stopping surveillance). The relationship between the TDF domains and themes provided a theoretical process describing factors impacting active surveillance adherence for men with low‐risk prostate cancer. Conclusions Men identified key factors impacting active surveillance adherence that provide opportunities for clinical implementation and practice improvement. Future efforts should focus on multi‐level interventions that foster trust in surveillance as treatment, emphasize quality of life benefits and enhance patients' interpersonal experiences while on surveillance to optimize adherence.

Abstract This study reports the incidence of chronic graft versus host disease (GvHD) in allogeneic hematopoietic stem cell transplant (alloHCT) recipients who received SARS-CoV2 vaccination. The overall rates of new and worsening chronic GvHD combined were 14%, with median time from vaccination to GVHD being approximately three to four weeks. A majority of the cases were of mild to moderate severity and primarily localized to either the skin, mouth, or joints. Prior chronic GVHD and recent transplant were associated with higher GVHD rates following COVID-19 vaccination. More prospective studies are needed to provide a definitive mechanism for the impact of SARS-CoV2 vaccination on alloHCT patients.

Objective(s): To elucidate the mechanism of Respiratory Syncytial Virus (RSV) infection and central neuronal disease and to understand the role of microglia in neuronal injuries during RSV infection.Materials and Methods: The effects of RSV and the cytokines produced by RSV-infected CHME-5 microglial cells on SY5Y neuronal cells were evaluated based on an in vitro Transwell coculture system. Five treatment groups were established in this study, including the normal control SY5Y group, RSV+SY5Y infection group, (cytokine+CHME-5)+SY5Y Transwell group, (RSV+CHME-5)+SY5Y Transwell group, and (RSV+cytokine+CHME-5)+SY5Y Transwell group. The morphological and physical alterations in SY5Y cells and their synapses were analyzed by confocal microscopy. The mRNA and protein expression levels of TLR3/RIG-I, as well as the expression of Hv1, in microglia were measured by qRT-PCR and Western blot assays. In addition, the apoptosis ratio of neuronal cells was determined by flow cytometry.Results: RSV infection activated the protein expression of Hv1 protein in microglia in vitro (p

Given that one of the major goals of the practice of Philosophy for Children (P4C) is the development of critical thinking skills (Sharp 1987/2018, pp. 4 6), an urgent question that emerged for one of the authors, who is of Chinese Heritage and a novice practitioner at a P4C summer camp (thinkingplayground.org), was whether this emphasis on critical thinking might make this practice incompatible with the fabric of Chinese culture. Filial piety (孝), which requires respect for one’s parents, elders, and ancestors is considered an important virtue in Asian culture, as is the preservation of harmony. But if one of the goals of P4C is to teach youngsters to courageously pursue reasoned dialogue, does this not set-up young Asians for serious conflict when they come face-to-face with positions that are articulated by elders, but which are ones to which they are diametrically opposed; a racist grandmother, for instance, or an uncle who insists that those at the Tiananmen Square uprising were nothing but hooligan’s. It is this question that we will explore in this presentation. In the process, we will come to the conclusion that, when positions seem irreconcilable, rather than continuing to pursue rigorous critical interchange that may do little other than escalate insult, the facilitator, rather, ought to move toward creating a deeper understanding of each position juxtaposed against its opposing view (a process that we refer to as ‘collaborative caring’), so as to produce side-by-side understanding, knowing that communal bonds have been maintained and, hence, that the opportunity for genuine reasoned collaborative inquiry on other issues and at future times remains open.

Asthma currently affects more than 339 million people worldwide. In the present preliminary study, we examined the efficacy of a new, inhalable soluble epoxide hydrolase inhibitor (sEHI), 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU), to attenuate airway inflammation, mucin secretion, and hyper-responsiveness (AHR) in an ovalbumin (OVA)-sensitized murine model. Male BALB/c mice were divided into phosphate-buffered saline (PBS), OVA, and OVA+TPPU (2- or 6-h) exposure groups. On days 0 and 14, the mice were administered PBS or sensitized to OVA in PBS. From days 26-38, seven challenge exposures were performed with 30 min inhalation of filtered air or OVA alone. In the OVA+TPPU groups, a 2- or 6-h TPPU inhalation preceded each 30-min OVA exposure. On day 39, pulmonary function tests (PFTs) were performed, and biological samples were collected. Lung tissues were used to semi-quantitatively evaluate the severity of inflammation and airway constriction and the volume of stored intracellular mucosubstances. Bronchoalveolar lavage (BAL) and blood samples were used to analyze regulatory lipid mediator profiles. Significantly (p < 0.05) attenuated alveolar, bronchiolar, and pleural inflammation; airway resistance and constriction; mucosubstance volume; and inflammatory lipid mediator levels were observed with OVA+TPPU relative to OVA alone. Cumulative findings indicated TPPU inhalation effectively inhibited inflammation, suppressed AHR, and prevented mucosubstance accumulation in the murine asthmatic model. Future studies should determine the pharmacokinetics (i.e., absorption, distribution, metabolism, and excretion) and pharmacodynamics (i.e., concentration/dose responses) of inhaled TPPU to explore its potential as an asthma-preventative or -rescue treatment.

A recent consensus guideline recommends migrating the therapeutic drug monitoring practice for intravenous vancomycin for the treatment of methicillin-resistant Staphylococcus aureus infection from the traditional trough-based approach to the Bayesian approach based on area under curve to improve clinical outcomes. To support the implementation of the new strategy for hospitals under Hospital Authority, Hong Kong, this study is being proposed to (1) estimate and validate a population pharmacokinetic model of intravenous vancomycin for local adults, (2) develop a web-based individual dose optimization application for clinical use, and (3) evaluate the performance of the application by comparing the treatment outcomes and clinical satisfaction against the traditional approach. 300 adult subjects prescribed with intravenous vancomycin and not on renal replacement therapy will be recruited for population pharmacokinetic model development and validation. Sex, age, body weight, serum creatinine level, intravenous vancomycin dosing records, serum vancomycin concentrations etc. will be collected from several electronic health record systems maintained by Hospital Authority. Parameter estimation will be performed using non-linear mixed-effect modeling techniques. The web-based individual dose optimization application is based on a previously reported application and is built using R and the package shiny. Data from another 50 subjects will be collected during the last three months of the study period and treated as informed by the developed application and compared against historical control for clinical outcomes. Since the study will incur extra blood-taking procedures from patients, informed consent is required. Other than that, recruited subjects should receive medical treatments as usual. Identifiable patient data will be available only to site investigators and clinicians in each hospital. The study protocol and informed consent forms have been approved by the Joint Chinese University of Hong Kong-New Territories East Cluster Clinical Research Ethics Committee (reference number: NTEC-2021-0215) and registered at the Chinese Clinical Trial Registry (registration number: ChiCTR2100048714).

Abstract Leptomeningeal metastasis is extremely rare in patients with ovarian cancer, but should be considered in patients presenting with neurologic deficits such as cauda equine syndrome. Given its poor prognosis and lack of data currently on management, additional studies are needed to optimize treatment regimens and improve outcomes.

Early detection and treatment of visual impairment diseases are critical and integral to combating avoidable blindness. To enable this, artificial intelligence–based disease identification approaches are vital for visual impairment diseases, especially for people living in areas with a few ophthalmologists. In this study, we demonstrated the identification of a large variety of visual impairment diseases using a coarse-to-fine approach. We designed a hierarchical deep learning network, which is composed of a family of multi-task & multi-label learning classifiers representing different levels of eye diseases derived from a predefined hierarchical eye disease taxonomy. A multi-level disease–guided loss function was proposed to learn the fine-grained variability of eye disease features. The proposed framework was trained for both ocular surface and retinal images, independently. The training dataset comprised 7,100 clinical images from 1,600 patients with 100 diseases. To show the feasibility of the proposed framework, we demonstrated eye disease identification on the first two levels of the eye disease taxonomy, namely 7 ocular diseases with 4 ocular surface diseases and 3 retinal fundus diseases in level 1 and 17 subclasses with 9 ocular surface diseases and 8 retinal fundus diseases in level 2. The proposed framework is flexible and extensible, which can be inherently trained on more levels with sufficient training data for each subtype diseases (e.g., the 17 classes of level 2 include 100 subtype diseases defined as level 3 diseases). The performance of the proposed framework was evaluated against 40 board-certified ophthalmologists on clinical cases with various visual impairment diseases and showed that the proposed framework had high sensitivity and specificity with the area under the receiver operating characteristic curve ranging from 0.743 to 0.989 in identifying all identified major causes of blindness. Further assessment of 4,670 cases in a tertiary eye center also demonstrated that the proposed framework achieved a high identification accuracy rate for different visual impairment diseases compared with that of human graders in a clinical setting. The proposed hierarchical deep learning framework would improve clinical practice in ophthalmology and broaden the scope of service available, especially for people living in areas with a few ophthalmologists.

Abstract Purpose To identify clinically relevant magnetic resonance imaging (MRI) features of different types of metastatic brain lesions, including standard anatomical, diffusion weighted imaging (DWI) and dynamic susceptibility contrast (DSC) perfusion MRI. Methods MRI imaging was retrospectively assessed on one hundred and fourteen (N = 114) brain metastases including breast (n = 27), non-small cell lung cancer (NSCLC, n = 43) and ‘other’ primary tumors (n = 44). Based on 114 patient’s MRI scans, a total of 346 individual contrast enhancing tumors were manually segmented. In addition to tumor volume, apparent diffusion coefficients (ADC) and relative cerebral blood volume (rCBV) measurements, an independent component analysis (ICA) was performed with raw DSC data in order to assess arterio-venous components and the volume of overlap (AVOL) relative to tumor volume, as well as time to peak (TTP) of T2* signal from each component. Results Results suggests non-breast or non-NSCLC (‘other’) tumors had higher volume compare to breast and NSCLC patients (p = 0.0056 and p = 0.0003, respectively). No differences in median ADC or rCBV were observed across tumor types; however, breast and NSCLC tumors had a significantly higher “arterial” proportion of the tumor volume as indicated by ICA (p = 0.0062 and p = 0.0018, respectively), while a higher “venous” proportion were prominent in breast tumors compared with NSCLC (p = 0.0027) and ‘other’ lesions (p = 0.0011). The AVOL component was positively related to rCBV in all groups, but no correlation was found for arterial and venous components with respect to rCBV values. Median time to peak of arterial and venous components were 8.4 s and 12.6 s, respectively (p

It is well-known that all progressive chronic kidney disease (CKD) is pathologically characterized by tubulointerstitial fibrosis process. Multiple studies have shown the critical role of inflammation and fibrosis in the development of CKD. Hence strategies that target inflammatory and fibrotic signaling pathways may provide promising opportunities to protect against renal fibrosis. Metformin has been used as the first-line glucose-lowering agent to treat patients with type 2 diabetes mellitus (T2DM) for over 50 years. Accumulating evidence suggests the potential for additional therapeutic applications of metformin, including mitigation of renal fibrosis. In this study, the anti-fibrotic effects of metformin independent of its glucose-lowering mechanism were examined in an adenine -induced mouse model of CKD. Expressions of inflammatory markers MCP-1, F4/80 and ICAM, fibrotic markers type IV collagen and fibronectin, and the cytokine TGF-β1 were increased in adenine-induced CKD when compared to control groups and significantly attenuated by metformin treatment. Moreover, treatment with metformin inhibited the phosphorylation of Smad3, ERK1/2, and P38 and was associated with activation of the AMP-activated protein kinase (AMPK) in the kidneys of adenine-treated mice. These results indicate that metformin attenuates adenine-induced renal fibrosis through inhibition of TGF-β1 signaling pathways and activation of AMPK, independent of its glucose-lowering action.

Dysregulated metabolism is a hallmark of cancer that manifests through alterations in bioenergetic and biosynthetic pathways to enable tumor cell proliferation and survival. Tumor cells exhibit high rates of glycolysis, a phenomenon known as the Warburg effect, and an increase in glutamine consumption to support the tricarboxylic acid (TCA) cycle. Renal cell carcinoma (RCC) tumors express high levels of glutaminase (GLS), the enzyme required for the first step in metabolic conversion of glutamine to glutamate and the entry of glutamine into the TCA cycle. We found that RCC cells are highly dependent on glutamine for proliferation, and this dependence strongly correlated with sensitivity to telaglenstat (CB-839), an investigational, first-in-class, selective, orally bioavailable GLS inhibitor. Metabolic profiling of RCC cell lines treated with telaglenastat revealed a decrease in glutamine consumption, which was concomitant with a decrease in the production of glutamate and other glutamine-derived metabolites, consistent with GLS inhibition. Treatment of RCC cells with signal transduction inhibitors everolimus (mTOR inhibitor) or cabozantinib (VEGFR/MET/AXL inhibitor) in combination with telaglenastat resulted in decreased consumption of both glucose and glutamine and synergistic anti-proliferative effects. Treatment of mice bearing Caki-1 RCC xenograft tumors with cabozantinib plus telaglenastat resulted in reduced tumor growth compared to either agent alone. Enhanced anti-tumor activity was also observed with the combination of everolimus plus telaglenastat. Collectively, our results demonstrate potent, synergistic, anti-tumor activity of telaglenastat plus signal transduction inhibitors cabozantinib or everolimus via a mechanism involving dual inhibition of glucose and glutamine consumption.

A highly reduced extrinsic pathway coagulation model (8 ODEs) under flow considered a thin 15-micron platelet layer where transport limitations were largely negligible (except for fibrinogen) and where cofactors (FVIIa, FV, FVIII) were not rate-limiting. By including thrombin feedback activation of FXI and the antithrombin-I activities of fibrin, the model accurately simulated measured fibrin formation and thrombin fluxes. Using this reduced model, we conducted 10,000 Monte Carlo (MC) simulations for ±50% variation of 5 plasma zymogens and 2 fibrin binding sites for thrombin. A sensitivity analysis of zymogen concentrations indicated that FIX activity most influenced thrombin generation, a result expected from hemophilia A and B. Averaging all MC simulations confirmed both the mean and standard deviation of measured fibrin generation on 1 tissue factor (TF) molecule per μm2. Across all simulations, free thrombin in the layer ranged from 20 to 300 nM (mean: 50 nM). The top 2% of simulations that produced maximal fibrin were dominated by conditions with low antithrombin-I activity (decreased weak and strong sites) and high FIX concentration. In contrast, the bottom 2% of simulations that produced minimal fibrin were dominated by low FIX and FX. The percent reduction of fibrin by an ideal FXIa inhibitor (FXI = 0) ranged from 71% fibrin reduction in the top 2% of MC simulations to only 34% fibrin reduction in the bottom 2% of MC simulations. Thus, the antithrombotic potency of FXIa inhibitors may vary depending on normal ranges of zymogen concentrations. This reduced model allowed efficient multivariable sensitivity analysis.

Early detection of vulnerable plaques is the critical step in the prevention of acute coronary events. Morphology, composition, and mechanical property of a coronary artery have been demonstrated to be the key characteristics for the identification of vulnerable plaques. Several intravascular multimodal imaging technologies providing co-registered simultaneous images have been developed and applied in clinical studies to improve the characterization of atherosclerosis. In this paper, the authors review the present system and probe designs of representative intravascular multimodal techniques. In addition, the scientific innovations, potential limitations, and future directions of these technologies are also discussed.

In order to evaluate the pharmacokinetics of recombinant bovine interferon-alpha (rBoIFN-α) in cattle, which has potential for its antiviral and immunomodulatory activities, 12 animals of 6-month age were classified into 4 groups (n=3) to receive rBoIFN-α through IV, IM or SC routes at a dose of 5.0×103 IU/kg. Serum rBoIFN-α titer was evaluated using cytopathic effect (CPE) inhibition bioassay. Then, the standard pharmacokinetic parameters were calculated using the DAS (Drug and statistics) software. The concentration-time profiles of serum rBoIFN-α following IM administration, SC administration and IV administration were characteristics of the 1-, 1-, and 2-compartment open models, respectively. After a single dose of IV administration, the drug rapidly dispersed and was rapidly eliminated from the body (T1/2α=0.15±0.02 h, T1/2=6.48±0.49 h). After IM and SC administrations, the drug is rapidly absorbed and slowly eliminated from the body (For IM administration, Tmax=6.12±0.32 h, T1/2=8.19±0.74 h) (For SC administration, Tmax=4.06±0.56 h, T1/2=7.29±0.55 h). The bioavailability of rBoIFN-α after IM administration is 53.74%, which is higher than the bioavailability of SC administration (27.96%). Therefore, the results showed that the drug administration effect can be preferably obtained following a single dose IM injection using the rBoIFN-α aqueous preparation. We hope that this study will provide valuable information for the clinical application of rBoIFN-α as an potential antiviral agent.

Abstract Background Over a third of multiple myeloma (MM) patients report clinical levels of depression during autologous stem cell transplant (ASCT) hospitalization. We report preliminary results from a randomized clinical trial investigating the effect of Programmed Environmental Illumination (PEI) of hospital rooms on depression. Methods Patients (N = 187) scheduled to receive an ASCT were assessed for eligibility. Those who met study eligibility criteria (n = 44) were randomly assigned to one of two PEI conditions involving delivery of either circadian active bright white light (BWL) or circadian inactive dim white light (DWL) throughout the room from 7 to 10 am daily during hospitalization. Patients completed the Center for Epidemiological Studies Depression Scale (CES‐D) prior to hospitalization, at days 2 and 7 post‐transplant, and on the third day of engraftment. Results General linear model analyses revealed no difference between the groups in CES‐D total score at baseline (P = 0.7859). A longitudinal linear mixed model analysis revealed a significant interaction between time of assessment and light condition [F(3,107) = 2.90; P = 0.0386; ɳ2 = 0.08)], indicating that PEI prevented the development of depression during hospitalization, with effects reaching significance by the third day of engraftment. At the third day of engraftment, 68.4% of the participants in the DWL comparison condition met the criteria for clinically significant depression compared to 42.1% in the BWL condition. Conclusion These findings demonstrate that PEI using BWL during MM ASCT hospitalization is effective in reducing the development of depression. Future studies should examine the mechanisms whereby PEI improves depression.

Abstract Background Myeloid cells are an abundant leukocyte in many types of tumors and contribute to immune evasion. Expression of the enzyme arginase 1 (Arg1) is a defining feature of immunosuppressive myeloid cells and leads to depletion of L-arginine, a nutrient required for T cell and natural killer (NK) cell proliferation. Here we use CB-1158, a potent and orally-bioavailable small-molecule inhibitor of arginase, to investigate the role of Arg1 in regulating anti-tumor immunity. Methods CB-1158 was tested for the ability to block myeloid cell-mediated inhibition of T cell proliferation in vitro, and for tumor growth inhibition in syngeneic mouse models of cancer as a single agent and in combination with other therapies. Tumors from animals treated with CB-1158 were profiled for changes in immune cell subsets, expression of immune-related genes, and cytokines. Human tumor tissue microarrays were probed for Arg1 expression by immunohistochemistry and immunofluorescence. Cancer patient plasma samples were assessed for Arg1 protein and L-arginine by ELISA and mass spectrometry, respectively. Results CB-1158 blocked myeloid cell-mediated suppression of T cell proliferation in vitro and reduced tumor growth in multiple mouse models of cancer, as a single agent and in combination with checkpoint blockade, adoptive T cell therapy, adoptive NK cell therapy, and the chemotherapy agent gemcitabine. Profiling of the tumor microenvironment revealed that CB-1158 increased tumor-infiltrating CD8+ T cells and NK cells, inflammatory cytokines, and expression of interferon-inducible genes. Patient tumor samples from multiple histologies expressed an abundance of tumor-infiltrating Arg1+ myeloid cells. Plasma samples from cancer patients exhibited elevated Arg1 and reduced L-arginine compared to healthy volunteers. Conclusions These results demonstrate that Arg1 is a key mediator of immune suppression and that inhibiting Arg1 with CB-1158 shifts the immune landscape toward a pro-inflammatory environment, blunting myeloid cell-mediated immune evasion and reducing tumor growth. Furthermore, our results suggest that arginase blockade by CB-1158 may be an effective therapy in multiple types of cancer and combining CB-1158 with standard-of-care chemotherapy or other immunotherapies may yield improved clinical responses.

Although rare, both central nervous system and pericardial involvement of myeloma have been well described in the literature. Their simultaneous occurrence in relapsed disease, however, has not been previously reported. This case describes a 54-year-old female who was treated for high-risk multiple myeloma with multiregimen chemotherapy and allogeneic hematopoietic stem cell transplantation. Four months after transplant, she was found to have relapsed disease manifesting as an extraosseous, intracranial plasmacytoma and simultaneous malignant pericardial effusion. Her disease characteristics, treatment course, radiologic and pathologic findings are described in detail, and we review the previous literature to determine the various aspects of her disease that may have contributed to her aggressive clinical course.

Bone fracture is a very common body injury. The healing process is physiologically complex, involving both biological and mechanical aspects. Following a fracture, cell migration, cell/tissue differentiation, tissue synthesis, and cytokine and growth factor release occur, regulated by the mechanical environment. Over the past decade, bone healing simulation and modeling has been employed to understand its details and mechanisms, to investigate specific clinical questions, and to design healing strategies. The goal of this effort is to review the history and the most recent work in bone healing simulations with an emphasis on both biological and mechanical properties. Therefore, we provide a brief review of the biology of bone fracture repair, followed by an outline of the key growth factors and mechanical factors influencing it. We then compare different methodologies of bone healing simulation, including conceptual modeling (qualitative modeling of bone healing to understand the general mechanisms), biological modeling (considering only the biological factors and processes), and mechanobiological modeling (considering both biological aspects and mechanical environment). Finally we evaluate different components and clinical applications of bone healing simulation such as mechanical stimuli, phases of bone healing, and angiogenesis.

Herein, we reported an Escherichia coli-based expression and purification method of recombinant porcine interferon alpha (rPoIFN-α). PoIFN-α coding sequence was cloned into pMD18-T vector and then subcloned into pET-32a (+) vector using standard recombinant DNA techniques and the resulting plasmid was transformed into BL21(DE3) competent cells. After induction with isopropyl-β-D-1-thiogalactopyranoside (IPTG), rPoIFN-α was purified from the supernatant of the bacteria lysate using a simple two-step chromatography process consisting of a Ni2+ affinity chromatography and a DEAE anion exchange chromatography. rPoIFN-α was purified to >95% homogeneity with a yield of 48 mg/L of culture. It has isoelectic point of 6.09 and bacterial endotoxin was less than 1 EU/mg. N-terminal amino acid sequence and the peptide map digested by trypsin provided additional evidence for the authenticity of rPoIFN-α. The biological activity of rPoIFN-α was 1.1×106 IU/ mL in HEp-2/ Vesicular Stomatitis Virus (VSV) titration system and its specific activity reached to 1.0×106 IU/mg. In conclusion, we obtained high-level expression of a soluble form of bioactive rPoIFN-α by using pET-32a (+) prokaryotic expression system.

We report the first published case of acute pancreatitis secondary to disseminated nocardiosis in a hematopoietic stem cell transplant (HSCT) recipient on chronic immunosuppression for graft-versus-host disease (GVHD). Nocardiosis in the HSCT population is relatively rare, and has not yet been described in haploidentical HSCT recipients. Our patient is a 28-year-old male with a history of haploidentical HSCT and GVHD of the skin and lung who was admitted to the hospital with acute pancreatitis. The workup for the etiology of his pancreatitis was initially unrevealing. He subsequently developed worsening sepsis and respiratory failure despite broad spectrum antimicrobials. After multiple bronchoscopies and pancreatic fluid sampling, he was found to have disseminated nocardiosis with Nocardia cyriacigeorgia.

During thrombosis, thrombin generates fibrin, however fibrin reversibly binds thrombin with low affinity E-domain sites (KD = 2.8 μM) and high affinity γ'-fibrin sites (KD = 0.1 μM). For blood clotting on collagen/tissue factor (1 TF-molecule/μm2) at 200 s-1 wall shear rate, high μM-levels of intraclot thrombin suggest robust prothrombin penetration into clots. Setting intraclot zymogen concentrations to plasma levels (and neglecting cofactor rate limitations) allowed the linearization of 7 Michaelis-Menton reactions between 6 species to simulate intraclot generation of: Factors FXa (via TF/VIIa or FIXa), FIXa (via TF/FVIIa or FXIa), thrombin, fibrin, and FXIa. This reduced model [7 rates, 2 KD's, enzyme half-lives~1 min] predicted the measured clot elution rate of thrombin-antithrombin (TAT) and fragment F1.2 in the presence and absence of the fibrin inhibitor Gly-Pro-Arg-Pro. To predict intraclot fibrin reaching 30 mg/mL by 15 min, the model required fibrinogen penetration into the clot to be strongly diffusion-limited (actual rate/ideal rate = 0.05). The model required free thrombin in the clot (~100 nM) to have an elution half-life of ~2 sec, consistent with measured albumin elution, with most thrombin (>99%) being fibrin-bound. Thrombin-feedback activation of FXIa became prominent and reached 5 pM FXIa at >500 sec in the simulation, consistent with anti-FXIa experiments. In predicting intrathrombus thrombin and fibrin during 15-min microfluidic experiments, the model revealed "cascade amplification" from 30 pM levels of intrinsic tenase to 15 nM prothrombinase to 15 μM thrombin to 90 μM fibrin. Especially useful for multiscale simulation, this reduced model predicts thrombin and fibrin co-regulation during thrombosis under flow.

While treatment with surgery, radiotherapy and/or chemotherapy may prolong life for patients with glioblastoma, recurrence is inevitable. What is still being discovered is how much these treatments and recurrence of disease affect the molecular profiles of these tumors and how these tumors adapt to withstand these treatment pressures. Understanding such changes will uncover pathways used by the tumor to evade destruction and will elucidate new targets for treatment development. Nineteen matched pre-treatment and post-treatment glioblastoma tumors were subjected to gene expression profiling (Fluidigm, TaqMan assays), MGMT promoter methylation analysis (pyrosequencing) and protein expression analysis of the DNA repair pathways, known to be involved in temozolomide resistance (immunohistochemistry). Gene expression profiling to molecularly subtype tumors revealed that 26% of recurrent post-treatment specimens did not match their primary diagnostic specimen subtype. Post-treatment specimens had molecular changes which correlated with known resistance mechanisms including increased expression of APEX1 (p < 0.05) and altered MGMT methylation status. In addition, genes associated with immune suppression, invasion and aggression (GPNMB, CCL5, and KLRC1) and polarization toward an M2 phenotype (CD163 and MSR1) were up-regulated in post-treatment tumors, demonstrating an overall change in the tumor microenvironment favoring aggressive tumor growth and disease recurrence. This was confirmed by in vitro studies that determined that glioma cell migration was enhanced in the presence of M2 polarized macrophage conditioned media. Further, M2 macrophage-modulated migration was markedly enhanced in post-treatment (temozolomide resistant) glioma cells. These findings highlight the ability of glioblastomas to evade not only the toxic onslaught of therapy but also to evade the immune system suggesting that immune-altering therapies may be of value in treating this terrible disease.

Microfluidic thrombosis assays allow the control of anticoagulation, hemodynamics, pharmacology, and procoagulant surfaces containing collagen ± tissue factor (TF). With corn trypsin inhibitor (CTI) ranging from low (1–4 μg/mL) to high levels (40–60 μg/mL), the function of Factor XIIa (FXIIa) can be modulated in the presence of low or high surface TF. With high CTI and no collagen/TF in the assay, no thrombin is generated during 15-min microfluidic perfusion. At low CTI (no TF), the generation of FXIa leads to fibrin polymerization at ~300 s after the initiation of flow over collagen, an onset time shortened at zero CTI and prolonged at high CTI. The engagement of FXIa was difficult to observe for clotting on high TF surfaces due to the dominance of the extrinsic pathway. Low TF surfaces allowed observable crosstalk between extrinsic pathway generation of thrombin and thrombin-mediated activation of FXIa, a feedback detected at >5 min and attenuated with polyphosphate inhibitor. From thrombin-antithrombin immunoassay of the effluent of blood flowing over collagen/TF, the majority of thrombin was found captured on intrathrombus fibrin. Additionally, extreme shear rates (>10,000 s−1) can generate massive von Willebrand Factor fibers that capture FXIIa and FXIa to drive fibrin generation, an event that facilitates VWF fiber dissolution under fibrinolytic conditions. Finally, we found that occlusive sterile thrombi subjected to pressure drops >70 mm-Hg/mm-clots have interstitial stresses sufficient to drive NETosis. These microfluidic studies highlight the interaction of contact pathway factors with the extrinsic pathway, platelet polyphosphate, VWF fibers, and potentially shear-induced NETs.

Despite optimal control of hyperglycaemia, hypertension, and dyslipidaemia, the number of patients with diabetic nephropathy (DN) continues to grow. Strategies to target various signaling pathways to prevent DN have been intensively investigated in animal models and many have been proved to be promising. However, targeting these pathways once kidney disease is established, remain unsatisfactory. The clinical scenario is that patients with diabetes mellitus often present with established kidney damage and need effective treatments to repair and reverse the kidney damage. In this studies, eNOS-/- mice were administered with streptozotocin to induce diabetes. At 24 weeks, at which time we have previously demonstrated albuminuria and pathological changes of diabetic nephropathy, mice were randomised to receive TRAM34 subcutaneously, a highly selective inhibitor of potassium channel KCa3.1 or DMSO (vehicle) for a further 14 weeks. Albuminuria was assessed, inflammatory markers (CD68, F4/80) and extracellular matrix deposition (type I collagen and fibronectin) in the kidneys were examined. The results clearly demonstrate that TRAM34 reduced albuminuria, decreased inflammatory markers and reversed extracellular matrix deposition in kidneys via inhibition of the TGF-β1 signaling pathway. These results indicate that KCa3.1 blockade effectively reverses established diabetic nephropathy in this rodent model and provides a basis for progressing to human studies.

AIMS/HYPOTHESIS:Developmental programming induced by maternal obesity influences the development of chronic disease in offspring. In the present study, we aimed to determine whether maternal obesity exaggerates obesity-related kidney disease. METHODS:Female C57BL/6 mice were fed high-fat diet (HFD) for six weeks prior to mating, during gestation and lactation. Male offspring were weaned to normal chow or HFD. At postnatal Week 8, HFD-fed offspring were administered one dose streptozotocin (STZ, 100 mg/kg i.p.) or vehicle control. Metabolic parameters and renal functional and structural changes were observed at postnatal Week 32. RESULTS:HFD-fed offspring had increased adiposity, glucose intolerance and hyperlipidaemia, associated with increased albuminuria and serum creatinine levels. Their kidneys displayed structural changes with increased levels of fibrotic, inflammatory and oxidative stress markers. STZ administration did not potentiate the renal effects of HFD. Though maternal obesity had a sustained effect on serum creatinine and oxidative stress markers in lean offspring, the renal consequences of maternal obesity were overwhelmed by the powerful effect of diet-induced obesity. CONCLUSION:Maternal obesity portends significant risks for metabolic and renal health in adult offspring. However, diet-induced obesity is an overwhelming and potent stimulus for the development of CKD that is not potentiated by maternal obesity.

This article presents results of a study of the impact of political dynamics on group deliberations of issues presented in the short film The Response. We selected four groups of 18-22 year-old participants based on political views, engagement, and efficacy (liberal, conservative, and two mixed groups), and asked them to view and discuss issues presented in The Response related to the combatant status review tribunals held at Guantanamo Bay. We found the groups with mixed political views had higher quality discussions of the issues and a better understanding of the issues post-discussion – in particular the tension between national security versus individual rights and of the nature of the tribunals. We also found a significant number of conservative group members became more conservative in their views as a result of their discussion. We discuss implications for secondary and post-secondary education as well as for political polarization overall in society.

Multiple rodent models have been used to study diabetic kidney disease (DKD). The purpose of the present study was to compare models of diabetes and obesity-induced metabolic syndrome and determine differences in renal outcomes. C57BL/6 male mice were fed either normal chow or high fat diet (HFD). At postnatal week 8, chow-fed mice were randomly assigned to low-dose streptozotocin (STZ, 55 mg/kg/day, five consecutive days) or vehicle control, whereas HFD-fed mice were given either one high-dose of STZ (100 mg/kg) or vehicle control. Intraperitoneal glucose tolerance tests were performed at Week 14, 20 and 30. Urinary albumin to creatinine ratio (ACR) and serum creatinine were measured, and renal structure was assessed using Periodic Acid Schiff (PAS) staining at Week 32. Results showed that chow-fed mice exposed to five doses of STZ resembled type 1 diabetes mellitus with a lean phenotype, hyperglycaemia, microalbuminuria and increased serum creatinine levels. Their kidneys demonstrated moderate tubular injury with evidence of tubular dilatation and glycogenated nuclear inclusion bodies. HFD-fed mice resembled metabolic syndrome as they were obese with dyslipidaemia, insulin resistance, and significantly impaired glucose tolerance. One dose STZ, in addition to HFD, did not worsen metabolic features (including fasting glucose, non esterified fatty acid, and triglyceride levels). There were significant increases in urinary ACR and serum creatinine levels, and renal structural changes were predominantly related to interstitial vacuolation and tubular dilatation in HFD-fed mice.

The activity of transforming growth factor-β1 (TGF-β1) is regulated by its conversion from the latent to the active form. We have previously shown that the conversion is at least in part mediated by the cationic-independent mannose 6-phosphate receptor (CI-M6PR), as the CI-M6PR inhibitor, PXS-25 has anti-fibrotic properties in human kidney tubular (HK-2) cells under high glucose conditions. However, its clinical use is limited by low bioavailability. Our aim was to determine the effects of PXS64, a pro-drug of PXS25, in in vitro and in vivo models of renal fibrosis. HK-2 cells were exposed to latent TGFβ1+/- PXS64 for 48 hours. The mRNA and protein levels of pro-fibrotic and pro-inflammatory markers were determined. A 7 day unilateral ureteric obstruction (UUO) model was used and the following experimental groups were studied: (i) Sham operated, (ii) UUO, (iii) UUO + telmisartan (iv) UUO + PSX64. HK-2 cells exposed to PXS64 reduced TGFβ mediated effects on collagen IV, fibronectin, macrophage chemotactic protein-1 (MCP-1) and phospho-smad2 protein expression, consistent with inhibition of the conversion of latent to active TGF-β1. PXS 64 treated UUO mice had a lower tubulointerstitial fibrosis index, collagen IV and fibronectin protein and mRNA expression when compared to untreated UUO mice. In addition, these animals had lower MCP-1 mRNA expression, reduced inflammarory cell infiltrate, as indicated by fewer CD45, F4/80 positive cells, and reduced phospho-Smad2 protein expression when compared to untreated UUO animals. Our data demonstrates that PSX64 is an effective anti-fibrotic agent by inhibiting the activation of latent TGF-β1.