Biomedical subjects

Naim Akhtar Khan

Publications and source records attributed to Naim Akhtar Khan.

8 recordsLinked to original sources

PubMed2005

n-3 PUFAs modulate T-cell activation via protein kinase C-alpha and -epsilon and the NF-kappaB signaling pathway.

We elucidated the mechanisms of action of two n-3 PUFAs, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), in Jurkat T-cells. Both DHA and EPA were principally incorporated into phospholipids in the following order: phosphatidylcholine < phosphatidylethanolamine < phosphatidylinositol/phosphatidylserine. Furthermore, two isoforms of phospholipase A(2) (i.e., calcium-dependent and calcium-independent) were implicated in the release of DHA and EPA, respectively, during activation of these cells. The two fatty acids inhibited the phorbol 12-myristate 13-acetate (PMA)-induced plasma membrane translocation of protein kinase C (PKC)-alpha and -epsilon. The two n-3 PUFAs also inhibited the nuclear translocation of nuclear factor kappaB (NF-kappaB) and the transcription of the interleukin-2 (IL-2) gene in PMA-activated Jurkat T-cells. Together, these results demonstrate that DHA and EPA, being released by two isoforms of phospholipase A(2), modulate IL-2 gene expression by exerting their action on two PKC isoforms and NF-kappaB in Jurkat T-cells.

Cell Membrane↗

PubMed2004

Thapsigargin-stimulated MAP kinase phosphorylation via CRAC channels and PLD activation: inhibitory action of docosahexaenoic acid.

This study was conducted on human Jurkat T-cells to investigate the role of depletion of intracellular Ca(2+) stores in the phosphorylation of two mitogen-activated protein kinases (MAPKs), i.e. extracellular signal-regulated kinase (ERK) 1 and ERK2, and their modulation by a polyunsaturated fatty acid, docosahexaenoic acid (DHA). We observed that thapsigargin (TG) stimulated MAPK activation by store-operated calcium (SOC) influx via opening of calcium release-activated calcium (CRAC) channels as tyrphostin-A9, a CRAC channel blocker, and two SOC influx inhibitors, econazole and SKF-96365, diminished the action of the former. TG-stimulated ERK1/ERK2 phosphorylation was also diminished in buffer containing EGTA, a calcium chelator, further suggesting the implication of calcium influx in MAPK activation in these cells. Moreover, TG stimulated the production of diacylglycerol (DAG) by activating phospholipase D (PLD) as propranolol (PROP) (a PLD inhibitor), but not U73122 (a phospholipase C inhibitor), inhibited TG-evoked DAG production in these cells. DAG production and protein kinase C (PKC) activation were involved upstream of MAPK activation as PROP and GF109203X, a PKC inhibitor, abolished the action of TG on ERK1/ERK2 phosphorylation. Furthermore, DHA seems to act by inhibiting PKC activation as this fatty acid diminished TG- and phorbol 12-myristate 13-acetate-induced ERK1/ERK2 phosphorylation in these cells. Together these results suggest that Ca(2+) influx via CRAC channels is implicated in PLD/PKC/MAPK activation which may be a target of physiological agents such as DHA.

Calcium↗

PubMed2004

n-3 fatty acids modulate T-cell calcium signaling in obese macrosomic rats.

OBJECTIVE: We investigated the effects of a diet containing EPAX-7010, rich in PUFAs such as eicosapentaenoic acid [20:5(n-3)] and docosahexaenoic acid [22:6(n-3)], i.e., a PUFA/EPAX regimen, on T-cell activation in diabetic pregnant rats and their obese pups. RESEARCH METHODS AND PROCEDURES: Mild hyperglycemia in pregnant rats was induced by intraperitoneal injection of streptozotocin on Day 5 of gestation. T-cell blastogenesis was assayed by using (3)H-thymidine, whereas intracellular free calcium concentrations ([Ca(2+)]i) were measured by using Fura-2 in diabetic pregnant rats and their obese offspring. RESULTS: Concavalin-A-stimulated T-cell proliferation was decreased in both pregnant diabetic rats and their obese pups as compared with control animals. Feeding the PUFA/EPAX diet restored T-cell proliferation in both groups of animals. We also employed ionomycin, which at 50 nM opens calcium channels, and thapsigargin (TG), which recruits [Ca(2+)]i from endoplasmic reticulum pool. We observed that ionomycin-induced increases in [Ca(2+)]i in T-cells of diabetic mothers and obese offspring were greater than in those of control rats. Furthermore, feeding PUFA/EPAX diet diminished significantly the ionomycin-evoked rise in [Ca(2+)]i in diabetic and obese animals. TG-induced increases in [Ca(2+)]i in T-cells of diabetic pregnant rats and their obese offspring were greater than in those of control rats. The feeding of the experimental diet significantly curtailed the TG-evoked increases in [Ca(2+)]i in both diabetic and obese rats. DISCUSSION: Together, these observations provide evidence that T-cell activation and T-cell calcium signaling are altered during gestational diabetes and macrosomia. Hence, dietary fish oils, particularly eicosapentaenoic acid and docosahexaenoic acid, may restore these T-cell abnormalities.

Animals↗

PubMed2003

Docosahexaenoic acid and other fatty acids induce a decrease in pHi in Jurkat T-cells.

1. Docosahexaenoic acid (DHA) induced rapid (t1/2=33 s) and dose-dependent decreases in pHi in BCECF-loaded human (Jurkat) T-cells. Addition of 5-(N,N-dimethyl)-amiloride, an inhibitor of Na+/H+ exchanger, prolonged DHA-induced acidification as a function of time, indicating that the exchanger is implicated in pHi recovery. 2. Other fatty acids like oleic acid, arachidonic acid, eicosapentaenoic acid, but not palmitic acid, also induced a fall in pHi in these cells. 3. To assess the role of calcium in the DHA-induced acidification, we conducted experiments in Ca2+-free (0% Ca2+) and Ca2+-containing (100% Ca2+) buffer. We observed that there was no difference in the degree of DHA-induced transient acidification in both the experimental conditions, though pHi recovery was faster in 0% Ca2+ medium than that in 100% Ca2+ medium. 4. In the presence of BAPTA, a calcium chelator, a rapid recovery of DHA-induced acidosis was observed. Furthermore, addition of CaCl2 into 0% Ca2+ medium curtailed DHA-evoked rapid pHi recovery. In 0% Ca2+ medium, containing BAPTA, DHA did not evoke increases in [Ca2+]i, though this fatty acid still induced a rapid acidification in these cells. These observations suggest that calcium is implicated in the long-lasting DHA-induced acidosis. 5. DHA-induced rapid acidification may be due to its deprotonation in the plasma membrane (flip-flop model), as suggested by the following observations: (1) DHA with a -COOH group induced intracellular acidification, but this fatty acid with a -COOCH3 group failed to do so, and (2) DHA, but not propionic acid, -induced acidification was completely reversed by addition of fatty acid-free bovine serum albumin in these cells. 6. These results suggest that DHA induces acidosis via deprotonation and Ca2+ mobilization in human T-cells.

Docosahexaenoic Acids↗

PubMed2002

Implication of three isoforms of PLA(2) in human T-cell proliferation.

We observed that human (Jurkat) T-cells constitutively expressed the mRNA, encoding for the four isoforms of phospholipase A(2) (PLA(2)), i.e. two secretory (type IB and type V), and two cytosolic (type IV, Ca(2+)-dependent and type VI, Ca(2+)-independent). In order to assess whether these PLA(2) isoforms are active, we labeled Jurkat T-cells with [(3)H]arachidonic acid ([(3)H]AA) and determined its release into the extracellular medium in the presence of phorbol 12-myristate 13-acetate (PMA) and ionomycin. The three PLA(2) isoforms seem functional as aristolochic acid and bromoenol lactone (BEL), the respective inhibitors of type IB/type V and type VI PLA(2)s, significantly inhibited the release of free [(3)H]AA. On the other hand, arachidonyl trifluoromethyl ketone (AACOCF(3)), an inhibitor of type IV PLA(2), failed to curtail significantly the release of free [(3)H]AA into the extracellular medium. We assessed the implication of these PLA(2) isoforms in transcription of the interleukin-2 (IL-2) gene, involved in T-cell proliferation. Hence, aristolochic acid and BEL, but not AACOCF(3), significantly inhibited the PMA and ionomycin-induced induction of mRNA of IL-2. Similarly, aristolochic acid and BEL, but not AACOCF(3), significantly inhibited the PMA and ionomycin-induced secretion of IL-2 in the culture supernatants. Together these results suggest that human Jurkat T-cells possess two secretory and two cytosolic PLA(2) isoforms and only three of them (type IB, type V and type VI) are implicated in T-cell proliferation.

Arachidonic Acid↗

PubMed2002

Impaired lipoprotein metabolism in obese offspring of streptozotocin-induced diabetic rats.

The time course of changes in lipoprotein metabolism of obese offspring of mildly diabetic rats was studied with respect to serum lipoprotein composition as well as LCAT and tissue lipoprotein lipase (LPL) activities. Mild hyperglycemia in pregnant rats was induced by intraperitoneal injection of streptozotocin on day 5 of gestation. Control pregnant rats were injected with citrate buffer. At birth, obese pups had higher serum glucose, insulin, and lipoprotein (VLDL, LDL-HDL1, HDL(2-3)) levels than control pups. After 1 mon of life, all of these parameters in obese rats became similar to those of controls. However, LCAT, adipose tissue LPL, and hepatic triacylglycerol lipase activities were high. At 2 mon of age, VLDL-TAG levels were higher in obese females than in controls. By the age of 3 mon, obese offspring had developed insulin resistance with hyperglycemia, hyperinsulinemia, and higher serum lipoprotein concentrations. Indeed, qualitative abnormalities of lipoproteins were seen and were typical of obese and diabetic human beings. Fetal hyperinsulinemia should be considered as a risk factor for later metabolic diseases, including dyslipoproteinemia.

Adipose Tissue↗

PubMed2002

Eicosapentaenoic acid and docosahexaenoic acid modulate MAP kinase enzyme activity in human T-cells.

In order to investigate the implication of docosahexacnoic acid (DHA) and eicosapentaenoic acid (EPA) in T signalling, we assessed their effects on the activation of two mitogen activated protein (MAP) kinases, i.e. extracellularly-regulated kinases 1 and 2 (ERK1/ERK2) in Jurkat T-cells. The n-3 polyunsaturated fatty acids (PUFAs) alone failed to induce MAP kinase (MAPK) enzyme activity. To elucidate whether DHA and EPA act via protein kinase C (PKC) dependent and independent pathways, we employed their respective activators, i.e. phorbol 12-myristate 13-acetate (PMA) and antiCD3 antibodies. We observed that U0126, an inhibitor of MAPK kinase-ERK kinase 1/2 (MEK1/2), abolished the actions of these two agents on MAPK activation, suggesting that they act upstream of MEK1/2. Further EPA and DHA diminished both the PMA- and antiCD3 antibodies-induced enzyme activity of ERK1/ERK2 in Jurkat T-cells. Interestingly, okadaic acid (OA), a phosphatase inhibitor seems to act downstream of MEK1/2 as U0126 failed to inhibit the OA-induced MAPK activation. It is noteworthy that EPA and DHA not only failed to curtail the OA-induced MAPK activity but also these n-3 PUFAs at 20 microM potentiated the action of OA. Therefore, EPA and DHA seem to modulate MAPK activation upstream and downstream of MEK1/2. On the hand, arachidonic acid, an n-6 PUFA potentiated the MAPK enzyme activity. In conclusion, our study shows that EPA and DHA may regulate T-cells functions by modulating MAPK enzyme activity.

Antibodies↗

PubMed2002

Role of three isoforms of phospholipase A2 in capacitative calcium influx in human T-cells.

The present study was conducted on human Jurkat T-cell lines in order to elucidate the role of phospholipase A2 in capacitative calcium entry. We have employed thapsigargin (TG) that induces increases in [Ca2+]i by emptying the calcium pool of endoplasmic reticulum, followed by capacitative calcium entry. We designed a Ca2+ free/Ca2+ reintroduction (CFCR) protocol for the experiments, conducted in Ca2+-free medium. By employing CFCR protocol, we observed that addition of exogenous arachidonic acid (AA) stimulated TG-induced capacitative calcium influx. The liberation of endogenous AA and its autocrine action seems to be implicated during TG-induced capacitative calcium influx: TG potentiates the induction of constitutively expressed mRNA of four PLA2 isoforms (type 1B, IV, V, VI), the inhibitors of the three PLA2 isotypes (type 1B, V, VI) inhibit TG-induced release of [3H]AA into the extracellular medium, and finally, these PLA2 inhibitors do curtail TG-stimulated capacitative calcium entry in these cells. These results suggest that stimulation of three isoforms of PLA2 by thapsigargin liberates free AA that, in turn, induces capacitative calcium influx in human T-cells.

Arachidonic Acid↗