A Review of Cases of Marijuana and Violence.

Marijuana is the most consumed illicit drug in the world, with over 192 million users. Due to the current legalization push of marijuana in the United States, there has been a lack of oversight regarding its public health policies, as marijuana advocates downplay the drug's negative effects. This paper's approach is from a public health perspective, focusing specifically on the cases of violence amongst some marijuana users. Here, we present 14 cases of violence with chronic marijuana users that highlight reoccurring consequences of: marijuana induced paranoia (exaggerated, unfounded distrust) and marijuana induced psychosis (radical personality change, loss of contact with reality). When individuals suffering from pre-existing medical conditions use marijuana in an attempt to alleviate their symptoms, ultimately this worsens their conditions over time. Although marijuana effects depend on the individual's endocannabinoid receptors (which control behavioral functions, like aggression) and the potency level of tetrahydrocannabinol (THC) in the drug, scientifically documented links between certain marijuana users and violence do exist. Wider public awareness of the risks and side effects of marijuana, as well as a more prudent health policy, and government agency monitoring of the drug's composition, creation, and distribution, are needed and recommended.

Channel catfish (Ictalurus punctatus) leukocytes express estrogen receptor isoforms ERα and ERß2 and are functionally modulated by estrogens.

Estrogens are recognized as modulators of immune responses in mammals and teleosts. While it is known that the effects of estrogens are mediated via leukocyte-specific estrogen receptors (ERs) in humans and mice, leucocyte-specific estrogen receptor expression and the effects of estrogens on this cell population is less explored and poorly understood in teleosts. Here in, we verify that channel catfish (Ictalurus punctaus) leukocytes express ERα and ERß2. Transcripts of these isoforms were detected in tissue-associated leukocyte populations by PCR, but ERß2 was rarely detected in PBLs. Expression of these receptors was temporally regulated in PBLs following polyclonal activation by concanavalin A, lipopolysaccharide or alloantigen based on evaluation by quantitative and end-point PCR. Examination of long-term leukocyte cell lines demonstrated that these receptors are differentially expressed depending on leukocyte lineage and phenotype. Expression of ERs was also temporally dynamic in some leukocyte lineages and may reflect stage of cell maturity. Estrogens affect the responsiveness of channel catfish peripheral blood leukocytes (PBLs) to mitogens in vitro. Similarly, bactericidal activity and phorbol 12-myristate 13-acetate induced respiratory burst was modulated by 17ß-estradiol. These actions were blocked by the pure ER antagonist ICI 182780 indicating that response is, in part, mediated via ERα. In summary, estrogen receptors are expressed in channel catfish leukocytes and participate in the regulation of the immune response. This is the first time leukocyte lineage expression has been reported in teleost cell lines.

Signaling proteins are represented in tissue fluid/lymph from soft tissues of normal human legs at concentrations different from serum.

BACKGROUND: The mobile intercellular fluid flowing to and in the lymphatics contains filtered plasma products and substances synthesized and excreted by tissue cells. Among them are signaling proteins such as cytokines, chemokines, enzymes, and growth factors. They act locally in autocrine and paracrine systems regulating cell metabolism, proliferation, and formation of the ground matrix. They play an immunoregulatory role in infections, wound healing, and tumor cell growth. METHODS AND RESULTS: In this study we measured the concentration of selected cytokines, chemokines, tissue enzymes, and growth factors in tissue fluid/lymph drained from normal human leg soft tissues. Legs exposed to infections and trauma often result in development of lymphedema. Lymph was drained from superficial calf lymphatics using microsurgical techniques. Our studies showed generally higher concentrations of cytokines, chemokines, enzymes, and growth factors in lymph than in serum. The total protein L/S ratio was 0.22, whereas that of various lymph signaling proteins ranged between 1 and 10. CONCLUSIONS: This indicates that in addition to proteins filtered from blood, local cells contribute to lymph concentration by own production, depending on the actual cell requirement. Moreover, there were major individual differences of lymph levels with simultaneous stable serum levels. This suggests existence of a local autonomous regulatory humoral mechanism in tissues, not reflected in serum.

Identification of two IgD+ B cell populations in channel catfish, Ictalurus punctatus.

Channel catfish Ictalurus punctatus express two Ig isotypes: IgM and IgD. Although catfish IgM has been extensively studied at the functional and structural levels, much less is known about IgD. In this study, IgM(+)/IgD(+) and IgM(-)/IgD(+) catfish B cell populations were identified through the use of anti-IgM and anti-IgD mAbs. Catfish IgM(+)/IgD(+) B cells are small and agranular. In contrast, IgM(-)/IgD(+) B cells are larger and exhibit a plasmablast morphology. The use of cell sorting, flow cytometry, and RT-PCR demonstrated that IgD(+) B cell expression varies among individuals. For example, some catfish have <5% IgM(-)/IgD(+) B cells in their PBLs, whereas in others the IgM(-)/IgD(+) B cell population can represent as much as 72%. Furthermore, IgD expressed by IgM(-)/IgD(+) B cells preferentially associates with IgL σ. Comparatively, IgM(+)/IgD(+) B cells can express any of the four catfish IgL isotypes. Also, transfection studies show that IgD functions as a typical BCR, because Igδ-chains associate with CD79a and CD79b molecules, and all membrane IgD transcripts from sorted IgM(-)/IgD(+) B cells contain viable VDJ rearrangements, with no bias in family member usage. Interestingly, all secreted IgD transcripts from IgM(+)/IgD(+) and IgM(-)/IgD(+) B cells were V-less and began with a leader spliced to Cδ1. Importantly, transfection of catfish clonal B cells demonstrated that this leader mediated IgD secretion. Together, these findings imply that catfish IgM(-)/IgD(+) B cells likely expand in response to certain pathogens and that the catfish IgD Fc-region, as has been suggested for human IgD, may function as a pattern recognition molecule.

Expression of alternatively spliced CD45 isoforms by channel catfish clonal T and B cells is dependent on activation state of the cell and regulated by protein synthesis and degradation.

In mammals, expression of the three alternatively spliced exons of the tyrosine phosphatase CD45 is regulated by the developmental and activation state of the cell. In comparison, the channel catfish, Ictalurus punctatus, CD45 homolog contains 18 functional alternatively spliced exons. Since very little is known about CD45 regulation in ectothermic vertebrates, this study examines the regulation of catfish CD45 mRNA isoform expression in clonal T and B cells in response to stimulation. Results show that mitogenic stimulation using catfish serum or concanavalin A induced expression of mRNAs for small CD45 isoforms, and isoform message expression was growth curve dependent, i.e. cells in logarithmic phase express message for smaller CD45 isoforms, whereas stationary phase cells express message for longer CD45 isoforms. In addition, cells treated with the protein synthesis inhibitor cycloheximide expressed message for longer CD45 isoforms, and treatment with lactacystin, which blocks protein degradation, rescued smaller isoform message expression. Collectively these data suggested that expression of CD45 isoforms, in catfish, at least at the mRNA level, is "constitutively dynamic" and highly dependent on extracellular stimuli.

An expanded self-antigen peptidome is carried by the human lymph as compared to the plasma.

BACKGROUND: The pre-nodal afferent lymph is the fluid which directly derives from the extracellular milieu from every parenchymal organ and, as it continues to circulate between the cells, it collects products deriving from the organ metabolism/catabolism. A comprehensive qualitative and quantitative investigation of the self-antigenic repertoire transported by the human lymph is still missing. METHODOLOGY/PRINCIPAL FINDINGS: A major difference between lymph and plasma could be visualized by FPLC and 2D gel in the amount of low molecular weight products corresponding to peptide fragments. Naturally processed peptides in normal pre-nodal human lymph were then fractionated by HPLC and characterized by multidimensional mass spectrometry. Analysis of more then 300 sequences identified self-peptides derived from both intracellular and extracellular proteins revealing the variety of catabolic products transported by human lymph. Quantitative analysis established that at least some of these peptides are present in the circulating lymph in nanomolar concentration. CONCLUSIONS/SIGNIFICANCE: The peptidome, generated by physiological tissue catabolism and transported by the pre-nodal lymph, is in addition to the self-peptidome generated in endosomal compartment. Unlike self antigen processed by local or nodal APC, which mostly produce epitopes constrained by the endosomal processing activity, self antigens present in the lymph could derived from a wider variety of processing pathways; including caspases, involved in cellular apoptosis, and ADAM and other metalloproteinases involved in surface receptor editing, cytokines processing and matrix remodeling. Altogether, expanding the tissue-specific self-repertoire available for the maintenance of immunological tolerance.

Surgical approaches to postobstetrical perineal body defects (rectovaginal fistula and chronic third and fourth-degree lacerations).

Rectovaginal fistulas and chronic anal sphincter lacerations are uncommon complications that are most often secondary to vaginal delivery, gynecologic surgery, and inflammatory bowel disease. In this chapter, we will review the pertinent anatomy, focusing on the 6 structures that should be considered during the repair and surgical techniques to promote restoration on normal anatomy and function. Key concepts include a tension-free repair, meticulous hemostasis, and postoperative bowel management.

Channel catfish soluble FcmuR binds conserved linear epitopes present on Cmu3 and Cmu4.

A linear epitope on catfish IgM has been identified as the docking site for the catfish soluble FcmuR (IpFcRI). Western blot analyses and latex bead binding assays identified the consensus octapeptide motif FxCxVxHE located at the second cysteine that forms the intrachain disulfide bond of the catfish Cmu3 and Cmu4 immunolglobulin (Ig) domains as the IpFcRI binding sites. Furthermore, molecular modeling of catfish Cmu3 and Cmu4 confirmed that the octapeptide in both of these domains is accessible for IpFcRI interactions. In addition, since this octapeptide motif is also found in other vertebrate Ig domains, IpFcRI binding to Ig heavy (H) and light (L) chains from rainbow trout, chicken, mouse, rabbit, and goat were examined by Western blot analyses and latex bead binding assays. IpFcRI readily bound reduced rainbow trout (Igmu), chicken (Ignu), mouse (Igmu, Iggamma1, Iggamma2a, Iggamma2b, and Igalpha), rabbit (Igmu and Iggamma) and goat (Iggamma) IgH chains, and mouse Igkappa and Iglambda, and chicken Iglambda IgL chains. IpFcRI also bound mouse IgM, IgA and IgG subclasses when examined under native conditions.

Immunoglobulin D enhances immune surveillance by activating antimicrobial, proinflammatory and B cell-stimulating programs in basophils.

Immunoglobulin D (IgD) is an enigmatic antibody isotype that mature B cells express together with IgM through alternative RNA splicing. Here we report active T cell-dependent and T cell-independent IgM-to-IgD class switching in B cells of the human upper respiratory mucosa. This process required activation-induced cytidine deaminase (AID) and generated local and circulating IgD-producing plasmablasts reactive to respiratory bacteria. Circulating IgD bound to basophils through a calcium-mobilizing receptor that induced antimicrobial, opsonizing, inflammatory and B cell-stimulating factors, including cathelicidin, interleukin 1 (IL-1), IL-4 and B cell-activating factor (BAFF), after IgD crosslinking. By showing dysregulation of IgD class-switched B cells and 'IgD-armed' basophils in autoinflammatory syndromes with periodic fever, our data indicate that IgD orchestrates an ancestral surveillance system at the interface between immunity and inflammation.

Genomic organization of the channel catfish CD45 functional gene and CD45 pseudogenes.

CD45 is a transmembrane protein tyrosine phosphatase, which in mammals plays an important role in T and B cell receptor and cytokine signaling. Recently, a catfish cDNA was shown to contain all characteristic CD45 features: an alternatively spliced amino-terminus, a cysteine-rich region, three fibronectin domains, a transmembrane region, and two phosphotyrosine phosphatase domains. However, analyses of CD45 cDNAs from various catfish lymphoid cell lines demonstrated that catfish CD45 is unique in that it contains a large number of alternatively spliced exons. Sequence analyses of cDNAs derived from the catfish clonal B cell line 3B11 indicated that this cell line expresses up to 13 alternatively spliced exons. Furthermore, sequence similarity among the alternatively spliced exons suggested duplication events. To establish the exact number and organization of alternatively spliced exons, a bacterial artificial chromosome library was screened, and the catfish functional CD45 gene plus six CD45 pseudogenes were sequenced. The catfish functional CD45 gene spans 37 kb and contains 49 exons. In comparison, the human and pufferfish CD45 genes consist of 34 and 30 exons, respectively. This difference in the otherwise structurally conserved catfish gene is due to the presence of 18 alternatively spliced exons that were likely derived through several duplication events. In addition, duplication events were also likely involved in generating the six pseudogenes, truncated at the 3' ends. A similarly 3' truncated CD45 pseudogene is also present in the pufferfish genome, suggesting that this specific CD45 gene duplication occurred before catfish and pufferfish diverged (approximately 400 million years ago).

Addictive disorders.

In conclusion, complex medical and psychiatric comorbidity is com-mon in individuals with substance use disorders. It is important to assess comorbidity because of the implications for prevention and treatment. Studies of the neurobiology of substance use and psychiatric disorders are accumulating rapidly and informing treatment development. Information about the prevention and treatment of infectious diseases and other medical conditions associated with substance use disorders also is growing, and it is important that patients are able to benefit from this. The articles in this issue provide state-of-the-art information about several issues related to comorbidity in substance use disorders.

Identification and characterization of a FasL-like protein and cDNAs encoding the channel catfish death-inducing signaling complex.

To elucidate cytolytic mechanisms in the channel catfish, lysates from catfish lymphoid and fibroblast cell lines were screened by Western blot analysis using a panel of antibodies reactive with components of the mammalian apoptotic pathway. Strong reactivity with three proteins (approximate Mr 70,000, 37,000, and 15,000) was seen using an antibody targeted to mammalian Fas ligand (FasL). The sizes of the two smaller proteins are consistent with their tentative designation as membrane-bound (37,000 Mr) and soluble (15,000 Mr) FasL. Treatments known to induce FasL in mammalian systems (e.g., PMA/calcium ionophore, UV-irradiation) induced expression of the 37,000- Mr protein in catfish T-cell lines. Moreover, expression of the 37,000- Mr protein in clonal T cells was up-regulated by increasing cell density. At the nucleotide level, homologues of Fas receptor (FasR), FADD, and caspase 8 were identified and characterized. These gene products likely constitute the teleost equivalent of the death-inducing signaling complex (DISC). FADD was constitutively expressed in all (T, B, macrophage, and fibroblast) cell lines examined as well as in peripheral blood lymphocytes (PBL), whereas FasR and caspase 8 were expressed in all cell lines except CCO, a FasL-positive fibroblast line. In contrast to FasL, expression of FasR and caspase 8 was inversely proportional to cell density. Collectively these studies identified four membrane-proximal proteins involved in the initiation of apoptosis in channel catfish and suggest that mechanisms of cell-mediated cytotoxicity in teleosts are similar to those used by mammals.

Human apoA-I/C-III/A-IV gene cluster transgenic rabbits: effects of a high-cholesterol diet.

We have generated transgenic rabbits that express the entire human apoA-I/C-III/A-IV gene cluster. As in humans, h-apoA-I and h-apoC-III were expressed in liver and intestine, whereas h-apoA-IV mRNA was detected in intestine only. Transgenic rabbits had significantly higher plasma total cholesterol, HDL-cholesterol and total phospholipid concentrations than non-transgenic littermates. In contrast to similar transgenic mice previously generated, which have gross hypertriglyceridemia, triglyceride concentrations were only moderately raised in transgenic rabbits. Plasma and HDL from transgenic rabbits were more effective than those from controls in promoting cholesterol efflux from cultured hepatoma cells. They had lower LCAT, lower CETP and higher PLTP activities than non-transgenic littermates. Cholesterol-feeding produced major increases in plasma lipids. The qualitative response to the diet was not modified by cluster expression. Human apoA-I concentration was halved by cholesterol-feeding, whereas h-apoC-III and h-apoA-IV concentrations were not significantly altered. Cholesterol efflux from hepatoma cells to plasma and HDL was not altered by the diet. Since lipoprotein metabolism of rabbits closely resembles that of humans, human apoA-I/C-III/A-IV transgenic rabbits may provide a reliable model for studies of the transcriptional regulation of the cluster, and for evaluating the effects of different agents on the expression of the three genes.

Reconstituted high-density lipoprotein attenuates organ injury and adhesion molecule expression in a rodent model of endotoxic shock.

The salutary effects of high-density lipoproteins (HDLs) in animal and human models of endotoxic shock have in the past been attributed to the ability of this lipoprotein to bind to lipopolysaccharide. However, the precise mechanisms for the protective effect of HDL are unclear. The first objective of this study was to determine the effects of HDLs on the organ injury and dysfunction associated with acute severe endotoxemia. Second, to gain insight into the mechanism of action of HDL, we also investigated the effect of HDLs on 1) the expression of P-selectin and intercellular adhesion molecule-1 in the kidneys of rats treated with endotoxin and 2) the rise in the plasma levels of tumor necrosis factor-alpha (TNF-alpha). Rats were given Escherichia coli lipopolysaccharide (6 mg/kg i.v.), pretreated with either vehicle (n = 9) or reconstituted HDL (rHDL; apolipoprotein A-I/phosphatidylcholine proteoliposomes, n = 10), and were monitored for 6 h. Here we report that rHDL attenuates the renal injury and dysfunction caused by endotoxin in the rat. In addition, rHDL reduced the degree of histological tissue injury in the lung, liver and intestine and attenuated the expression of P-selectin and intercellular adhesion molecule-1 in the renal glomerulus. Interestingly, pretreatment of rats with rHDL did not prevent the hypotension nor the rise in plasma levels of TNF-alpha (at 90 min) caused by endotoxin. Thus, rHDL reduces the organ injury/dysfunction, but does not affect the circulatory failure, nor the rise in plasma levels of TNF-alpha caused by endotoxin in the rat. We propose that the mechanisms of these beneficial effects of HDL may be related to direct inhibition of adhesion molecule expression.

Tibolone lowers high density lipoprotein cholesterol by increasing hepatic lipase activity but does not impair cholesterol efflux.

OBJECTIVE: Androgens and other drugs that reduce plasma concentrations of high density lipoprotein (HDL) cholesterol are often considered to be pro-atherogenic. Tibolone lowers HDL-cholesterol by 20% but the clinical significance of this effect is unknown. METHODS: In a randomized, double-blind study, 34 women received 2.5 mg tibolone daily and 34 women received placebo. Serum concentrations of lipids, lipoprotein subclasses and apolipoproteins, together with plasma activities of lipid transfer proteins and lipolytic enzymes and the capacity of plasma to induce cholesterol efflux from cultured cells, were measured. RESULTS: Compared to placebo, tibolone reduced serum concentrations of HDL-cholesterol (-14%), HDL phosphatidylcholine (-14%), apolipoprotein (apo)A-I (-12%), HDL subclasses lipoprotein (Lp)A-I (-20%), HDL-apoE (-16%), pre beta-LpA-I (-10%) and alpha-LpA-I (-12%) and increased hepatic lipase activity (+25%) and HDL sphingomyelin : phosphatidylcholine ratio (10.5%), but did not alter serum concentrations of HDL sphingomyelin, apoA-IV and LpA-I/A-II, lipoprotein lipase, the plasma activities of lecithin : cholesterol acyl transferase, cholesteryl ester transfer protein, phospholipid transfer protein or the plasma capacity to release cholesterol from cultured fibroblasts or Fu5AH hepatocytes. CONCLUSIONS: Tibolone lowers HDL-cholesterol in part by increasing hepatic lipase activity. Conservation of sphingomyelin and apoA-II in HDL, as well as cholesteryl ester transfer protein activity, preserves the capacity of plasma to release cholesterol, despite the lower concentrations of HDL-cholesterol. This may have important implications for the use of steroid effects on HDL concentrations as surrogates for atherosclerosis.

Immortal and mortal clonal lymphocyte lines from channel catfish: comparison of telomere length, telomerase activity, tumor suppressor and heat shock protein expression.

Channel catfish autonomous (immortal) and nonautonomous (mortal) leukocyte lines were phenotyped with respect to telomere length and the expression of telomerase, Hsp70 and p53, potentially important factors in cellular immortalization. The autonomous cells constitutively expressed telomerase whereas the nonautonomous cells expressed this activity only transiently. This observation, coupled with the low telomerase activity level seen in freshly isolated leukocytes, suggests that telomerase expression in catfish leukocytes is activation induced. In contrast both types of cell lines exhibited quite similar patterns of significantly shortened telomeres, suggesting that telomerase does not stabilize catfish telomeres until a critical short length is reached. Northern analyses indicated that, like telomerase, Hsp70 gene expression was constitutive in autonomous cells and transient in nonautonomous cells. In contrast, p53 mRNA levels appeared similarly low and noncycling in both long-term cultured types of catfish cells, regardless of the culture situation. Furthermore it was noted, by Western analyses, that both types of cells display multiple sized forms of p53 proteins. This latter observation implies that truncation of p53 protein is probably not directly involved in the in vitro immortalization process of channel catfish leukocytes.

Channel catfish cytotoxic cells: a mini-review.

The use of allogeneic and autologous lymphoid cell lines has facilitated studies of cytotoxic T lymphocytes (CTL) and natural killer (NK)-like cells in channel catfish. Naïve catfish leukocytes were shown to spontaneously kill allogeneic cells and virally-infected autologous cells without the need for prior sensitization, and allogeneic cytotoxic responses were greatly enhanced by in vitro alloantigen stimulation. Both catfish CTL and NK-like cells have been successfully cloned from these alloantigen-stimulated cultures, and represent the first cytotoxic cell lines derived from any ectothermic vertebrate. These cloned cytotoxic cells contain granules and likely induce apoptosis in sensitive targets via a putative perforin/granzyme mechanism. In addition, some catfish CTL clones may also kill targets by an additional mechanism, possibly by Fas/FasL-like interactions. Importantly, these cytotoxic cells do not express the marker for catfish nonspecific cytotoxic cells (NCCs), and thus represent cell types distinct from NCCs. The use of monoclonal antibodies against the catfish F and G immunoglobulin light chain isotypes revealed the presence of a putative Fc receptor for IgM (Fc mu R) on some catfish NK-like cells that appears to 'arm' these cells with surface IgM. In addition, a potentially important monoclonal antibody (CC41) developed against catfish NK-like cells was found to recognize an approximately 150kDa molecule on the surface of catfish cytotoxic cells. These studies clearly demonstrate that catfish possess an array of different cytotoxic cells. The availability of various cloned cytotoxic cell lines should enable unambiguous functional studies to be performed in ways not currently possible with any other fish species.