Effects of a putative antidepressant with a rapid onset of action in defeated mice with different coping strategies

There is evidence suggesting that stressful social events may result in depressive-like disorders, but the development of these disorders depend on the way in which people cope with stress. Although antidepressants are useful their drawback is a delay in the therapeutic effects, moreover not all the patients show an adequate response to this treatment. The aim of this study was to analyse the effect of RS 67333, which is a 5-HT(4) receptor partial agonist and a putative antidepressant which exhibits a rapid onset of action and to determine whether this drug reverses the behavioural and physiological effects that are generated by chronic defeat in subjects who manifest a more vulnerable profile in their response to stress. Male mice were exposed to defeat for 21 consecutive days using a sensorial contact model. After 18 days of defeat, 2 groups of subjects were established, active and passive, in accordance with the behaviour that was manifested during social confrontation, and drug treatment was initiated for 5 days. Finally, the animals were subjected to a forced swimming test (FST). The results revealed higher corticosterone levels in passive mice after the last defeat. Additionally, 3 days after the last defeat, they showed lower corticosterone levels and higher splenic IL-6 and TNF-α levels and hypothalamic GR mRNA levels when compared to their active and manipulated control counterparts. Passive mice had higher 5-HT(1A) receptor mRNA levels than the manipulated controls and a lower MR/GR ratio than active mice. Similar to stress, the drug increased hypothalamic GR mRNA levels, but it did not affect other measured physiological variables or social behaviour, which suggested that the mechanism of this drug is not the most adequate for reversing stress-induced effects in this model. Nevertheless, the treatment increased swimming and decreased immobility in the FST, suggesting an antidepressant potential for this drug.     

Impact of hepatitis C infection on long-term mortality of injecting drug users from 1990 to 2002: differences before and after HAART

OBJECTIVE: To assess the impact of HIV and hepatitis C virus (HCV) infection on long-term mortality in injecting drug users (IDU). DESIGN: Community-based prospective cohort study. METHODS: Mortality data from follow-up in clinical sites and the Mortality Registry by December 2002 were collected for 3247 IDU who attended three centres for voluntary counselling and testing for HIV/AIDS, HCV and hepatitis B virus (HBV) in 1990-1996. Mortality rates by Poisson regression were adjusting for age, sex, duration of drug use, education, HBV and calendar period (1990-1997 and 1998-2002). RESULTS: Overall, 11.2% were HIV/HCV negative, 43.7% positive only for HCV and 45.1% positive for both. During 26 772 person-years of follow-up, 585 deaths were detected (2.19/100 person-years). Before 1997, HIV/HCV-positive subjects had a five-fold increase in risk of death [relative risk (RR), 5.4; 95% confidence interval (CI), 2.5-11.4] compared with those negative for both; after 1997, a three-fold increase was observed (RR, 2.7; 95% CI, 1.7-4.2). Being HCV positive/HIV negative was not associated with an increase in the risk of death either before (RR, 1.3; 95% CI, 0.6-2.9) or after (RR, 1.2; 95% CI, 0.8-1.9) 1997 compared with HCV/HIV negative. While increases in mortality were seen in those HCV/HIV negative (RR, 1.6; 95% CI, 0.7-3.7) and those only positive for HCV (RR, 1.5; 95% CI, 1.0-2.1), a 20% reduction among coinfected IDUs was observed after 1997 (interaction P = 0.033). CONCLUSIONS: HCV/HIV coinfection has had a large impact on mortality in IDU. After 1997, mortality increased in HIV negative/HCV positive subjects and decreased in HIV positive/HCV positive.     

Antitumor efficacy and reduced toxicity using an anti-CD137 Probody therapeutic

Costimulation via CD137 (4-1BB) enhances antitumor immunity mediated by cytotoxic T lymphocytes. Anti-CD137 agonist antibodies elicit mild liver inflammation in mice, and the maximum tolerated dose of Urelumab, an anti-human CD137 agonist monoclonal antibody, in the clinic was defined by liver inflammation–related side effects. A protease-activated prodrug form of the anti-mouse CD137 agonist antibody 1D8 (1D8 Probody therapeutic, Pb-Tx) was constructed and found to be selectively activated in the tumor microenvironment. This construct, which encompasses a protease-cleavable linker holding in place a peptide that masks the antigen binding site, exerted antitumor effects comparable to the unmodified antibody but did not result in liver inflammation. Moreover, it efficaciously synergized with both PD-1 blockade and adoptive T-cell therapy. Surprisingly, minimal active Pb-Tx reached tumor-draining lymph nodes, and regional lymphadenectomy did not abrogate antitumor efficacy. By contrast, S1P receptor–dependent recirculation of T cells was absolutely required for efficacy. The preferential cleavage of the anti-CD137 Pb-Tx by tumor proteases offers multiple therapeutic opportunities, including neoadjuvant therapy, as shown by experiments in which the Pb-Tx is given prior to surgery to avoid spontaneous metastases.

Immunotherapy based on blocking T-cell coinhibitory receptors has revolutionized cancer clinical management with the advent of anti–PD-(L)1 and anti–CTLA-4 checkpoint inhibitors (1). Immunostimulatory agonist antibodies targeting costimulatory targets such as CD137 (4-1BB), OX40, ICOS, GITR, or CD27 are lagging behind in clinical development for a variety of reasons (2, 3).In the case of agonist anti-CD137 monoclonal antibodies (mAb), the immunotherapeutic effects in mouse models are impressive (4), especially when combined with anti–PD-(L)1 (5, 6), interleukin-12 (7), or adoptive T-cell therapy (ACT) (8, 9). The mechanism of action mainly relies on CD8 T-cell costimulation and reinvigoration of dysfunctional tumor-infiltrating lymphocytes (8, 10).In the clinic, the strong CD137 agonist Urelumab underwent several clinical trials (11) but was limited to subtherapeutic doses by frequent and serious liver inflammation above 0.3 mg/kg (11). Another anti-CD137 mAb, Utomilumab (12), shows weak intrinsic agonist activity and could be safely dosed up to 10 mg/kg but without consistent evidence for clinical activity as a monotherapy (13).Many attempts are ongoing in the clinic to safely target CD137 costimulation to the tumor while preserving liver safety (14, 15). These include bispecific antibodies (16) and tumor-targeted constructs containing trimeric natural ligand (CD137L) (17). Here, we report the immune and therapeutic effects of an agonist anti-CD137 antibody prodrug called a Probody therapeutic (Pb-Tx) (18, 19). The term Probody is a United States–registered trademark of CytomX Therapeutics to refer to a new class of recombinant, proteolytically activated antibody prodrugs. The anti-CD137 Pb-Tx is designed to be activated to exert agonist activity on CD137 only in the tumor when cleaved by tumor-associated proteases.     

A Novel Mouse Model of Endometriosis Mimics Human Phenotype and Reveals Insights into the Inflammatory Contribution of Shed Endometrium

Endometriosis is an estrogen-dependent inflammatory disorder characterized by the presence of endometrial tissue outside the uterine cavity. Patients experience chronic pelvic pain and infertility, with the most likely origin of the tissue deposits (lesions) being endometrial fragments shed at menses. Menstruation is an inflammatory process associated with a dramatic increase in inflammatory mediators and tissue-resident immune cells. In the present study, we developed and validated a mouse model of endometriosis using syngeneic menstrual endometrial tissue introduced into the peritoneum of immunocompetent mice. We demonstrate the establishment of endometriotic lesions that exhibit similarities to those recovered from patients undergoing laparoscopy. Specifically, in both cases, lesions had epithelial (cytokeratin⁺) and stromal (vimentin/CD10⁺) cell compartments with a well-developed vasculature (CD31⁺ endothelial cells). Expression of estrogen receptor β was increased in lesions compared with the peritoneum or eutopic endometrium. By performing experiments using mice with green fluorescent protein–labeled macrophages (MacGreen) in reciprocal transfers with wild-type mice, we obtained evidence that macrophages present in the peritoneum and in menses endometrium can contribute to the inflammatory microenvironment of the lesions. In summary, we developed a mouse model of endometriosis that exhibits similarities to human peritoneal lesions with respect to estrogen receptor expression, inflammation, and macrophage infiltration, providing an opportunity for further studies and the possible identification of novel therapies for this perplexing disorder.Endometriosis occurs in 6% to 10% of women of reproductive age,¹ is associated with chronic pelvic pain and infertility, and represents a huge socioeconomic burden.² The defining feature of the condition is the presence of endometrial tissue outside the uterine cavity, typically on the peritoneal wall³ or the surface of the ovary (endometriomas).⁴ Spontaneous endometriosis occurs only in humans and some primates,^5,6 where the luminal portion of the endometrium sheds at the end of the menstrual cycle. The primary cause of endometriosis is widely accepted to be the introduction of endometrial tissue into the peritoneal cavity via the fallopian tubes, a process known as retrograde menstruation.⁷ In captive baboons, the incidence of spontaneous endometriosis parallels the number of menstrual cycles experienced, providing a link between shedding of endometrial tissue and the onset of disease.⁸Current treatment strategies for peritoneal endometriosis are restricted to surgical excision of the lesions or suppression of ovarian function to mimic premature menopause. In up to 75% of cases, symptoms recur after surgery, with long-term ovarian suppression sometimes ineffective.⁹ Although it is acknowledged that there is an unmet clinical need for new nonsurgical treatments for endometriosis, their development depends on access to animal models that can recapitulate features of the human disorder.¹⁰ Nonhuman primates offer a physiologically relevant model of endometriosis, although their use is limited by cost and ethical concerns. Injecting female baboons with autologous menstrual endometrium into the peritoneal cavity has been shown to result in the development of endometriotic lesions.¹¹ This model has been used in longitudinal studies, allowing new insights into changes in gene expression in lesions as they become established.¹² In studies using the same model, expression of aromatase and the β isoform of estrogen receptor (ERβ) was found to be increased in the lesions, a finding in agreement with studies in women.^13,14 Rat and mouse models of endometriosis have also been developed, offering the advantage of lower cost and smaller animal size for testing potential therapies. The availability of mice genetically engineered to express fluorescent marker proteins^15,16 or targeted deletion of genes implicated in endometrial disease progression¹⁷ has made them an attractive alternative to primate studies. To date, studies using mice have usually involved the injection of human endometrial tissue into immunosuppressed mice¹⁸ or the introduction of small fragments of mouse uterine tissue that are sutured to the peritoneum or other sites (reviewed in Grummer¹⁹). Results obtained using both models have provided new insights into the origins of the disorder and its association with infertility²⁰ and have served as a platform for testing potential therapies, including those targeting angiogenesis in the lesions²¹; however, both models have limitations. For example, although human tissue injected either under the skin or into the peritoneal cavity can result in the formation of endometriosis-like lesions, these can grow only if the recipients lack the capacity to mount a tissue rejection response, eg, they are homozygous for the Prkdc^SCID mutation.¹⁸ Models using mice with an intact immune system allow for studies on the role(s) of inflammatory cytokines and immune cells,^22,23 but because mice do not exhibit spontaneous decidualization or menstruation, the uterine tissue introduced into the peritoneal cavity does not recapitulate the tissue microenvironment at the time of retrograde menstruation. Furthermore, in most of the autologous/syngeneic mouse models, uterine tissue is secured in the cavity after local abrasion and/or using sutures,^16,22 with both procedures likely to induce an artificial inflammatory response.In the present study, we set out to develop a mouse model of endometriosis that would combine the best features of the current baboon and mouse models. For this we used our recently validated mouse model of menstruation²⁴ as the source of syngeneic mouse menstrual endometrium and introduced this into the peritoneum of immunocompetent mice. To determine whether this approach could mimic the estrogenic and inflammatory features of human peritoneal lesions, we examined gene and protein expression in mice and patients and complemented these studies by undertaking reciprocal transfers between mice in which cells of the granulocyte/macrophage lineage were labeled with green fluorescent protein (GFP).¹⁵     

Inverted Takotsubo Cardiomyopathy and the Fundamental Diagnostic Role of Echocardiography

Takotsubo cardiomyopathy is characterized by the development of transient focal wall-motion abnormalities that involve the apical and midventricular segments, in the absence of obstructive coronary artery disease. A variant, inverted takotsubo cardiomyopathy, was described in 2010. We report 3 cases in which each patient''s transthoracic echocardiogram revealed the characteristic basal and midventricular segmental akinesis of this variant. This pattern is not associated with coronary artery distribution, and it therefore can be differentiated from coronary artery disease with the use of echocardiography, by evaluating the distribution and temporal changes of akinetic areas.Key words: Cardiomyopathies/diagnosis/physiopathology, echocardiography, takotsubo cardiomyopathy/diagnosis, ventricular dysfunction, left/diagnosisTakotsubo cardiomyopathy is a syndrome characterized by transient apical and midventricular akinesis that is typically precipitated by acute stress. A variant, with akinesis of the mid and basal left ventricular (LV) segments and sparing of the apex, has been called inverted (or atypical) takotsubo cardiomyopathy. We describe 3 cases of this variant in which the transthoracic echocardiograms (TTEs) and temporal evolution of the condition were diagnostic. Each patient was emergently admitted to Barnes-Jewish Hospital in St. Louis and examined there; the TTEs were reviewed by a visiting cardiologist (AM).     

Monitoring hepatitis C virus treatment rates in an Opioid Treatment Program:A longitudinal study

BACKGROUND Direct-acting antivirals(DAAs) are recommended for the treatment of hepatitis C virus(HCV) infection in patients treated with methadone or buprenorphine.AIM To assess HCV treatment rates in an Opioid Treatment Program(OTP).METHODS This longitudinal study included 501 patients(81.4% men, median age: 45 years; interquartile range: 39-50 years) enrolled in an OTP between October 2015 and September 2017. Patients were followed until September 2019. Data on sociodemographics, substance use, HCV infection, human immunodeficiency virus(HIV) infection and laboratory parameters were collected at entry. We analyzed medical records to evaluate HCV treatment. Kaplan-Meier methods and Cox regression models were used to analyze the DAA treatment uptake and to identify treatment predictors.RESULTS Prevalence of HCV and HIV infection was 70% and 34%, respectively. Among anti-HCV-positive(n = 336) patients, 47.2%, 41.3%, and 31.9% used alcohol,cannabis, and cocaine, respectively. HCV-RNA tests were positive in 233(69.3%) patients. Twentyeight patients(8.3%) cleared the infection, and 59/308(19.1%) had received interferon-based treatment regimens before 2015. Among 249 patients eligible, 111(44.6%) received DAAs. Treatment rates significantly increased over time from 7.8/100 person-years(p-y)(95%CI: 5.0-12.3) in 2015 to 18.9/100 p-y(95%CI: 11.7-30.3) in 2019. In a multivariate analysis, patients with HIV co-infection were twice as likely to receive DAAs(HR = 1.94, 95%CI: 1.21-3.12) than patients with HCV mono-infection. Current drug use was an independent risk factor for not receiving treatment against infection(HR = 0.48, 95%CI: 0.29-0.80).CONCLUSION HCV treatment is evolving in patients with HCV-HIV co-infection. Ongoing drug use while in an OTP might negatively impact the readiness to treat infection.     

Surfactant Protein D, a Marker of Lung Innate Immunity, Is Positively Associated With Insulin Sensitivity

OBJECTIVE

Impaired lung function and innate immunity have both attracted growing interest as a potentially novel risk factor for glucose intolerance, insulin resistance, and type 2 diabetes. We aimed to evaluate whether surfactant protein D (SP-D), a lung-derived innate immune protein, was behind these associations.

RESEARCH DESIGN AND METHODS

Serum SP-D was evaluated in four different cohorts. The cross-sectional associations between SP-D and metabolic and inflammatory parameters were evaluated in two cohorts, the cross-sectional relationship with lung function in one cohort, and the longitudinal effects of weight loss on fasting and circadian rhythm of serum SP-D and cortisol concentrations in one prospective cohort.

RESULTS

In the cross-sectional studies, serum SP-D concentration was significantly decreased in subjects with obesity and type 2 diabetes (P = 0.005) and was negatively associated with fasting and postload serum glucose. SP-D was also associated with A1C, serum lipids, insulin sensitivity, inflammatory parameters, and plasma insulinase activity. Smoking subjects with normal glucose tolerance, but not smoking patients with type 2 diabetes, showed significantly higher serum SP-D concentration than nonsmokers. Serum SP-D concentration correlated positively with end-tidal carbon dioxide tension (r = 0.54, P = 0.034). In the longitudinal study, fasting serum SP-D concentration decreased significantly after weight loss (P = 0.02). Moreover, the main components of cortisol and SP-D rhythms became synchronous after weight loss.

CONCLUSIONS

These findings suggest that lung innate immunity, as inferred from circulating SP-D concentrations, is at the cross-roads of inflammation, obesity, and insulin resistance.Impaired lung function has attracted growing interest in association with metabolic disorders (1–6). Decreased lung function has been proposed as a potential novel risk factor for glucose intolerance, insulin resistance, and type 2 diabetes (1–6). In prospective studies of middle-aged men and women without known lung disease, lower vital capacity predicted the subsequent development of type 2 diabetes. Lower forced vital capacity and forced expiratory volume in 1 s at baseline predicted hyperinsulinemia and estimated insulin resistance over 20 years of follow-up, independent of age, adiposity, and smoking (1).Possible mechanisms for the hypothesized link include direct effects of hypoxemia on glucose and insulin regulation (7), adverse early-life exposures and their effects on organ development (8), and lung-related inflammatory mediators and their effects on insulin signaling (9). In fact, nuclear factor interleukin-6, early growth response-1, and hypoxia-inducible factor-1 mediate inflammatory responses to chronic hypoxia in macrophages, pulmonary vascular endothelium, and smooth muscle (6,9). Cigarette smoking, an independent predictor of type 2 diabetes (10), provokes an inflammatory response (11) and is inversely associated with vital capacity. However, the link between lower vital capacity and diabetes risk was completely independent of cigarette exposure and was stronger in never-smokers (6).Reduced vital capacity is a common residual effect of lower respiratory tract infections, including those in childhood and infancy (8), that might provoke an inflammatory response. A reduced ability to sense and eradicate pathogens could thus cause frequent respiratory tract infections, reduced vital capacity, and chronic inflammation resulting in insulin resistance and type 2 diabetes (12). The total incidence rate of infections needing hospitalization in diabetic patients was 41/1,000 persons-years compared with 16/1,000 person-years of follow-up in the general population. Roughly half of the infections were severe lung infections, suggesting impaired lung immunity in patients with type 2 diabetes (13).Pulmonary surfactant is a complex mixture of lipids (90%) and proteins (5–10%) that constitutes the mobile liquid phase covering the large surface area of the alveolar epithelium. It maintains minimal surface tension within the lungs to avoid lung collapse during respiration. The innate immune system, by upregulating SP-D synthesis, can immediately respond to intrusion of foreign agents by helping to prevent further invasion (14). This recognition is important in the day-to-day physiology. Each day, we breathe >7,000 liters of air, laden with inorganic and organic particles and an array of microbes. Secreted primarily by alveolar epithelial type II pneumocytes, plasma SP-D appears to increase early in the clinical course of lung injury, and its concentration is thought to reflect pulmonary epithelial injury (15).Subtle deficiencies in proteins of the sensing arm of the innate immune system have been found to be associated with alterations of glucose metabolism. These deficiencies run in parallel with inflammation and impaired insulin action (16).We hypothesized that SP-D could be behind the association of lung function with impaired insulin action. For that reason, we aimed to evaluate SP-D according to metabolic and inflammatory parameters. As SP-D was associated with obesity status and impaired glucose metabolism, we evaluated the influence of weight loss on both fasting and circadian serum SP-D concentration. As glucocorticoids seem to regulate SP-D production in in vitro studies (17), we investigated the influence of circadian cortisol rhythm on serum SP-D concentration. Finally, we also studied the association of SP-D with lung function tests.     

A comparative morphologic and morphometric study about geniculate ganglion development in man and in rat

Abstract

Conclusions: Human–rat geniculate ganglion (GG) have multiple origins: (1) An initial proximity (20?μm) to the endocranial foramen of the IAM, suggests neural crest induction; and (2) The influence of epibranchial placodes: the tensor tympani muscle (TTM) and the otic apical coil.

Objectives: This study was undertaken to determine the comparative development of human–rat GG.

Materials and methods: A light microscopic study of the GG in human material obtained from spontaneous abortions at 9, 13, 14, 17, 18, and 30 weeks, and one neonate was done. This study examined Webster rat embryos and a post-natal series. Specimens were fixed in Bouin fluid, embedded in paraffin, cut, and stained with H&E. The histomorphometric data were obtained with image analysis software.

Results: The human fetus of 9 weeks presents two neuronal groups in the VII nerve: one near (20?μm) the IAM endocranial foramen, foraminal, and the other, tympanic. Neonate GG is located between the TTM and the cochlear apex (inwards). In the 16?day old rat embryo GG is placed within a canal containing the stapedial artery. In the adult rat the GG and the stapedial artery are placed within the IAM.