Nadeem Sh | University of the Punjab (original) (raw)

Papers by Nadeem Sh

European journal of medical research, Jan 28, 2015

Cytokines have always been of great interest due to their vast potential and participation in the... more Cytokines have always been of great interest due to their vast potential and participation in the progression and pathogenesis of various ailments. Interleukin-32 (IL-32) is a recently identified cytokine, whose gene is located on human chromosome 16 p13.3, with eight exons and six splice variants (IL-32¿ to IL-32¿). IL-32¿, the most abundant form, is secreted by different types of cells including T cells, natural killer (NK) cells, monocytes, endothelial cells and epithelial cells. It acts as a preferential mediator and effector of abnormal immune responses to multiple inflammatory and auto immune diseases including rheumatoid arthritis, chronic obstructive pulmonary disease (COPD), inflammatory bowel disease (IBD), etc. It was found to stimulate the induction of various chemokines, pro-inflammatory cytokines including IL-1ß, IL-6, IL-8, TNF-¿ and macrophage inflammatory protein-2 (MIP-2). Hence, IL-32 mediates the crucial interplay among immune system and body cells during pathoge...

Background: Thioacetamide (TAA) has been used extensively in the development of suitable animal m... more Background: Thioacetamide (TAA) has been used extensively in the development of suitable animal models of acute and chronic liver injury employing various doses, times and routes of its administration, particularly in drinking water due to its resemblance with human liver fibrosis and cirrhosis. The aim of this study was to investigate and compare hematological alteration during the acute and chronic liver inflammation. Methods: Acute Liver inflammation was in-duced in Wistar rats via intraperitoneal injection of thioacetamide and the animals were sacrificed 12 h after the TAA administration.Induction of chronic liver inflammation was performed by continuous administration of TAA in the drinking water (200 mg/L) during 18 weeks of experiment. After that all animals were sacrificed and Blood samples were collected for further analysis. Results: Single intra peritoneal injection of TAA (300 mg/kg B.W.) induced an acute condition with hematological changes including leukocytosis with marked neutrophilia (P = 0.0429), lymphopenia, thrombocytosis as well as increased hemoglobin concentration (P < 0.05) and decline of erythrocytic count (P = 0.0009). Eighteen weeks of uninterrupted supply of TAA (200 mg/L) in drinking water lead to chronic inflammation and the hematological alterations were leucopenia (P = 0.0197) accompanied with neutropenia and thrombocytopenia. Increase in RBCs (P = 0.0073) and Hb contents was also observed with a decline of red cell indices. Conclusion: Taken together these findings we can conclude that the animals respond differently under acute and chronic inflammatory condition with TAA administration. Leukocytosis with marked neutrophilia, thrombocytosis as well as increased hemoglobin concentration and decline of erythrocytic count were observed in acute while leucopenia accompanied with neutropenia and thrombocytopenia and increase in RBCs, Hb and Hct was also observed with a decline of other red cell indices during chronic phase.
Keywords: Haematology; Inflammation; Leucocytosis; Leucopenia; Neutrophilia; Neutropenia; Thrombocytosis and Thrombocytopenia

American journal of physiology. Gastrointestinal and liver physiology, 2006

In this work, we used two rat models, partial hepatectomy (PH) and CCl(4) administration, to stud... more In this work, we used two rat models, partial hepatectomy (PH) and CCl(4) administration, to study the changes in iron pathways in response to hepatic damage. Liver injury induced changes in the hepatic gene expression of hepcidin, hemojuvelin (Hjv), several other proteins of iron metabolism, and several cytokines such as IL-1beta, IL-6, TNF-alpha, and IFN-gamma. Hepcidin gene expression was upregulated between 4 and 8 h with a maximum up to 16 h after surgery. However, Hjv gene expression was downregulated at the same time. An early upregulation of hepcidin (3 h) and downregulation of Hjv gene expression was found after CCl(4) administration. Transferrin receptor 1 and ferritin H gene expression was upregulated, whereas ferroportin 1 gene expression was downregulated. Hepatic IL-6 gene expression was upregulated early after PH and reached maximum 8 h after the PH. In CCl(4)-induced liver injury, IL-6, IL-1beta, TNF-alpha, and IFN-gamma upregulation were found at the maximum 12 h after the administration of the toxin. Treatment of isolated rat hepatocytes with IL-6 and, to a lesser extent, with IL-1beta but not with TNF-alpha or IFN-gamma dose dependently upregulated hepcidin and downregulated Hjv gene expression. In hepatic damage, changes of the hepatic gene expression of the main proteins involved in iron metabolism may be induced by locally synthesized mediators.

Laboratory investigation; a journal of technical methods and pathology, 2006

Cytokine-induced neutrophil chemoattractant-1 (CINC-1), a member of the ELR + CXC subfamily [ELR ... more Cytokine-induced neutrophil chemoattractant-1 (CINC-1), a member of the ELR + CXC subfamily [ELR motif (glutamic acid-leucine-arginine) adjacent to the cysteine-X-cysteine (CXC) motif located at the N-terminus of the protein], is an acute-phase protein and its synthesis is induced by endogenous and exogenous pyrogens. However, there are no studies on the pyrogenic property of CINC-1. Therefore, the present study evaluates whether centrally administered CINC-1 promotes an integrated febrile response along with an increase in the prostaglandin (PG)E 2 content of the cerebrospinal fluid (CSF) of rats. The effects of antipyretic drugs on fever and on the PGE 2 content of the CSF as well as the effectiveness of a neutralizing anti-CINC-1 antibody on the fever induced by CINC-1 have also been investigated. Intracerebroventricular (i.c.v.) injection of CINC-1 induced a dosedependent bell-shaped rise on body temperature and increased PGE 2 concentration in the CSF of conscious rats. Injected into the preoptic area of the anterior hypothalamus (AH/POA) (i.h.), CINC-1 also induced a dose-dependent bell-shaped increase in body temperature along with a decrease on tail skin temperature. Indomethacin (INDO, 2 mg kg − 1 , i.p.) and ibuprofen (IBU, 10 mg kg − 1 , i.p.) markedly reduced the fever evoked by i.c.v. injection of CINC-1 (25 ng/site). Orally given celecoxib (5 mg kg − 1 , 30 min. before) abolished the fever induced by CINC-1 i.c.v. or i.h. (50 pg) injection. The antipyretic drugs also blocked the PGE 2 increase after CINC-1 i.c.v. injection. Co-injected anti-CINC antibody (10 ng/site) strongly reduced the febrile response induced by CINC-1 (50 pg/site) injected intrahypothalamically. This is the first time that centrally injected CINC-1 has been reported to act directly on the pyrogen-sensitive neurons of AH/POA, promoting a thermoregulatory response that seems to depend on other endogenous pyrogens synthesis and, as seen here, on PGE 2 .

Radiology, 2007

To prospectively analyze hepcidin, hemojuvelin, and ferroportin-1 expression after x-irradiation ... more To prospectively analyze hepcidin, hemojuvelin, and ferroportin-1 expression after x-irradiation of rat liver and isolated rat hepatocytes. The treatment of the rats and this study were approved by the local committee and the public authority on animal welfare. Rat livers in vivo and isolated rat hepatocytes in vitro were irradiated. The total number of rats in this study was 43. RNA extracted from livers (1, 3, 6, 12, 24, and 48 hours after irradiation) and from hepatocytes (1, 3, 6, 12, and 24 hours after irradiation) was analyzed with real-time polymerase chain reaction and Northern blot. Cytokines and prohepcidin in serum of irradiated rats were quantitatively detected with enzyme-linked immunosorbent assay. Sham-irradiated animals served as controls in all experiments. Differences between sham-irradiated and irradiated data groups were tested with analysis of variance and Dunnett post hoc test. In vivo, a significant radiation-induced increase of hepcidin (P=.034), interleukin (IL) 1beta (P=.008), IL-6 (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;.011), and tumor necrosis factor alpha (TNF-alpha) (P=.047) expression could be detected within the first 48 hours after irradiation. Expression of hemojuvelin (P=.008) and ferroportin-1 (P=.002) was significantly decreased. Serum iron levels were decreased because of irradiation (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;.058); prohepcidin serum levels were increased (P=.05). In rat hepatocytes in vitro, hepcidin RNA levels were significantly downregulated after irradiation (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;.001). Incubation of irradiated hepatocytes with IL-1beta, IL-6, or TNF-alpha led to upregulation of hepcidin expression in vitro up to 6 hours after irradiation, with subsequent significant downregulation for incubation with IL-1beta (P&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;.001). Hemojuvelin expression behaved in a way opposite to that of hepcidin. x-Irradiation of the liver induced changes of hepcidin gene expression that are probably induced by acute phase mediators produced within the liver itself.

Laboratory investigation; a journal of technical methods and pathology, 2007

Radiation research, 2008

The aim of the study was to analyze the effect of a single irradiation on chemokine gene expressi... more The aim of the study was to analyze the effect of a single irradiation on chemokine gene expression in the rat liver and in isolated rat hepatocytes. RNA extracted from livers and from hepatocytes within the first 48 h after irradiation was analyzed by real-time PCR and the Northern blot assay. The chemokine concentrations in the serum of irradiated rats were measured quantitatively by ELISA. A significant radiation-induced increase of CINC1, IP10, MCP1, MIP3alpha, MIP3beta, MIG and ITAC gene expression could be detected at the RNA level in the liver. CINC1, MCP1 and IP10 serum levels were significantly increased. In rat hepatocytes in vitro, only MIP3alpha showed a radiation-induced increase in expression, while CINC1, IP10, MIP3beta, MIG, MIP1alpha, ITAC and SDF1 RNA levels were significantly down-regulated. However, incubation of irradiated hepatocytes in vitro with either TNF-alpha, IL1beta, or IL6 plus TNF-alpha led to up-regulation of MCP1, IP10 and MCP1 or CINC1 and MIP3beta, respectively. Irradiation of the liver induces up-regulation of the genes of the main proinflammatory chemokines, probably through the action of locally synthesized proinflammatory cytokines. The reason for the lack of liver inflammation in this model has still to be clarified.

Laboratory investigation; a journal of technical methods and pathology, 2009

The liver and the spleen are the organs in which cellular material and aged erythrocytes are elim... more The liver and the spleen are the organs in which cellular material and aged erythrocytes are eliminated from the blood. Within the liver, Kupffer cells (KCs) are mainly responsible for this task, as such KCs have a pivotal role in iron metabolism. The aim of this study is to investigate the changes of hepatic gene expression in two models of KC phagocytosis. Gadolinium chloride (GD) or zymosan was injected intraperitoneally into rats and to endotoxin-resistant mice (C3H/HeJ). The animals were killed at different time points and their livers were immediately frozen in liquid nitrogen for RNA isolation and immunohistological studies. RNA was analyzed by real-time PCR and northern blot. Sera were used to measure transaminases, hepcidin and iron levels. The expression of iron metabolism genes, hepcidin, hemojuvelin (Hjv), ferroportin-1 (Fpn-1) and of the inflammatory cytokines IL-6, IL-1b, TNF-a and IFN-g was determined. Although phagocytosed material was detected in ED-1-and C1q-positive cells, no inflammatory cells were identified within the liver parenchyma. Serum levels of hepcidin, iron and transaminases did not differ from those of control animals. Both GD and zymosan induced an upregulation of hepcidin-gene expression in rat liver as early as 3 h, reaching a maximum 6 h after treatment. Hjv-and Fpn-1-gene expression was downregulated at the same time. IL-6 was by far the most induced acute-phase-cytokine in GD-and zymosan-treated livers, although IL-1b and TNF-a were also strongly upregulated by zymosan and to a lesser extent by GD. Similar results were obtained in the C3H/HeJ mouse strain excluding the possible role of contaminating endotoxin. This study shows that phagocytosis upregulates hepcidin-gene expression and downregulates Hjv-and Fpn-1-gene expression within the liver. These changes in iron-regulating-gene expression may be mediated by the locally produced acute-phase-cytokines.

Liver international : official journal of the International Association for the Study of the Liver, 2010

An acute-phase response is the systemic reaction of an organism to insult (e.g. infection, trauma... more An acute-phase response is the systemic reaction of an organism to insult (e.g. infection, trauma and burning). It represents the 'first line' of defence of the body to tissue-damaging attacks. In the present work, we used a rat model of an intra-muscular turpentine oil (TO) injection to analyse erythropoietin (EPO) gene expression changes in the liver, one of the main target organs of acute-phase cytokines. EPO began to increase in the serum of TO-treated animals 6 h after injection and reached a maximum at 24 h (125 AE 20 pg/ml). The detection of total RNA by polymerase chain reaction analysis showed that the levels of EPO gene expression in the liver were considerably increased between 2 and 12 h by up to 20-fold at the peak after TO administration, followed by a gradual decrease over the next 48 h, although the values remained significantly higher compared with the control group. In the kidney, after a sudden slight increase, the values declined progressively to 3.5-fold decrease at 12 h after the injection. In the liver, a parallel upregulation of the hypoxia-inducible factor-1 (HIF-1) a gene was observed (up to 4.7-fold increase), while HIF-2 a gene expression remained unaltered. On the other hand, the protein of both genes became detectable after the injection and increased progressively over 24 h, with a subsequent decline. These results suggest that EPO may be added to the increasing group of positive acute-phase proteins and the liver might represent the major source of the hormone under these conditions in the rat.

The American journal of pathology, 2010

Liver damage is a serious clinical complication of ␥-irradiation. We therefore exposed rats to si... more Liver damage is a serious clinical complication of ␥-irradiation. We therefore exposed rats to singledose ␥-irradiation (25 Gy) that was focused on the liver. Three to six hours after irradiation , an increased number of neutrophils (but not mononuclear phagocytes) was observed by immunohistochemistry to be attached to portal vessels between and around the portal (myo)fibroblasts (smooth muscle actin and Thy-1 ؉ cells). MCP-1/CCL2 staining was also detected in the portal vessel walls , including some cells of the portal area. CC-chemokine (MCP-1/CCL2 and MCP-3/CCL7) and CXC-chemokine (KC/CXCL1 , MIP-2/CXCL2 , and LIX/CXCL5) gene expression was significantly induced in total RNA from irradiated livers. In laser capture microdissected samples , an early (1 to 3 hours) up-regulation of CCL2 , CXCL1 , CXCL8, and CXCR2 gene expression was detected in the portal area but not in the parenchyma; with the exception of CXCL1 gene expression . In addition, treatment with an antibody against MCP-1/CCL2 before irradiation led to an increase in gene expression of interferon-␥ and IP-10/CXCL10 in liver tissue without influencing the recruitment of granulocytes. Indeed, the CCL2, CXCL1, CXCL2, and CXCL5 genes were strongly expressed and further up-regulated in liver (myo)fibroblasts after irradiation (8 Gy). Taken together, these results suggest that ␥-irradiation of the liver induces a transient accumulation of granulocytes within the portal area and that (myo)fibroblasts of the portal vessels may be one of the major sources of the chemokines involved in neutrophil recruitment. Moreover, inhibition of more than one chemokine (eg, CXCL1 and CXCL8) may be necessary to reduce leukocytes recruitment. (Am J Pathol 2010, 176:000 -000; DOI:

Cell and tissue research, 2010

Non-thyroidal illness is characterized by low triiodothyronine (T3) serum level under acute-phase... more Non-thyroidal illness is characterized by low triiodothyronine (T3) serum level under acute-phase conditions. We studied hepatic gene expression of the newly identified thyroid hormone receptor (TR) cofactor DOR/ TP53INP2 together with TRs in a rat model of aseptic abscesses induced by injecting intramuscular turpentine-oil into each hind limb. A fast (4-6 h) decrease in the serum level of free thyroxine and free T3 was observed. By immunohistology, abundant DOR protein expression was detected in the nuclei of hepatocytes and ED-1 + (mononuclear phagocytes), CK-19 + (biliary cells), and SMA + (mesenchymal cells of the portal tract) cells. DOR signal was reduced with a minimum at 6-12 h after the acute-phase reaction (APR). Immunohistology also showed a similar pattern of protein expression in TRα1 but without a significant change during APR. Transcripts specific for DOR, nuclear receptor corepressor 1 (NCoR-1), and TRβ1 were down-regulated with a minimum at 6-12 h, whereas expression for TRα1 and TRα2 was slightly and significantly up-regulated, respectively, with a maximum at 24 h after APR was initiated. In cultured hepatocytes, acute-phase cytokines interleukin-1β (IL-1β) and IL-6 down-regulated DOR and TRβ1 at the mRNA level. Moreover, gene expression of DOR and TRs (TRα1, TRα2, and TRβ1) was up-regulated in hepatocytes by adding T3 to the culture medium; this upregulation was almost completely blocked by treating the cells with IL-6. Thus, TRβ1, NCoR-1, and the recently identified DOR/TP53INP2 are abundantly expressed and down-regulated in liver cells during APR. Their downregulation is attributable to the decreased serum level of thyroid hormones and most probably also to the direct action of the main acute-phase cytokines.

Cell and tissue research, 2011

The "acute phase" is clinically characterized by homeostatic alterations such as somnolence, adin... more The "acute phase" is clinically characterized by homeostatic alterations such as somnolence, adinamia, fever, muscular weakness, and leukocytosis. Dramatic changes in iron metabolism are observed under acutephase conditions. Rats were administered turpentine oil (TO) intramuscularly to induce a sterile abscess and killed at various time points. Tissue iron content in the liver and brain increased progressively after TO administration. Immunohistology revealed an abundant expression of transferrin receptor-1 (TfR1) in the membrane and cytoplasm of the liver cells, in contrast to almost only nuclear expression of TfR1 in brain tissue. The expression of TfR1 increased at the protein and RNA levels in both organs. Gene expression of hepcidin, ferritin-H, iron-regulatory protein-1, and heme oxygenase-1 was also upregulated, whereas that of hemojuvelin, ferroportin-1, and the hemochromatosis gene was significantly downregulated at the same time points in both the brain and the liver at the RNA level. However, in contrast to observations in the liver, gene expression of the main acute-phase cytokine (interleukin-6) in the brain was significantly upregulated. In vitro experiments revealed TfR1 membranous protein expression in the liver cells, whereas nuclear and cytoplasmic TfR1 protein was detectable in brain cells. During the non-bacterial acute phase, iron content in the liver and brain increased together with the expression of TfR1. The iron metabolism proteins were regulated in a way similar to that observed in the liver, possibly by locally produced acute-phase cytokines. The significance of the presence of TfR1 in the nucleus of the brain cells has to be clarified.

Laboratory investigation; a journal of technical methods and pathology, 2012

Liver is the central organ of iron metabolism. During acute-phase-response (APR), serum iron conc... more Liver is the central organ of iron metabolism. During acute-phase-response (APR), serum iron concentration rapidly decreases. The current study aimed to compare expression and localization of iron transport protein ferroportin-1 (Fpn-1) and of other iron import proteins after experimental tissue damage induced by injecting turpentine oil in the hind limbs of rats and mice. Serum and spleen iron concentration decreased with an increase in total liver, cytoplasmic and nuclear iron concentration. In liver, mRNA amount of Fpn-1, Fpn-1a, Fpn-1b, HFE, hemojuvelin (HJV) and hephaestin (heph) genes showed a rapid decrease. Hepcidin, divalent metal transporter-1 (DMT-1), transferrin (Tf) and Tf-receptor-1 (TfR1), TfR-2 (TfR2) gene expression was increased. Western blot analysis of liver tissue lysate confirmed the changes observed at mRNA level. In spleen, a rapid decrease in gene expression of Fpn-1, Fpn-1a, Fpn-1b, DMT-1, Tf, TfR1 and TfR2, and an increase in hepcidin was observed. Immunohistochemistry of DMT-1 and TfR2 were mainly detected in the nucleus of rat liver and spleen, whereas TfR1 was clearly localized in the plasma membrane. Fpn-1 was mostly found in the nuclei of liver cells, whereas in spleen, the protein was mainly detected in the cell membrane. Western blot analysis of liver fractions confirmed immunohistochemical results. In livers of wild-type mice, gene expression of Fpn-1, Fpn-1a and Fpn-1b was downregulated, whereas hepcidin gene expression was increased. In contrast, these changes were less pronounced in IL-6ko-mice. Cytokine (IL-6, IL-1b and TNF-a) treatment of rat hepatocytes showed a downregulation of Fpn-1, Fpn-1a and Fpn-1b, and upregulation of hepcidin gene expression. Moreover, western blot analysis of cell lysate of IL-6-treated hepatocytes detected, as expected, an increase of a2-macroglobulin (positive acute-phase protein), whereas albumin (negative acute-phase protein) and Fpn-1 were downregulated. Our results demonstrate that liver behaves as a 'sponge' for iron under acute-phase conditions, and Fpn-1 behaves as a negative acute-phase protein in rat hepatocytes mainly, but not exclusively, because of the effect of IL-6. These changes could explain iron retention in the cytoplasm and in the nucleus of hepatocytes during APR.

International journal of clinical and experimental pathology, 2013