Major Cognitive Changes and Micrographia following Globus Pallidus Infarct.

Importance. Globus pallidus (GP) lesions are well known to cause motor deficits but are less commonly-and perhaps not conclusively-associated with cognitive problems. Observations. We present a 45-year-old male with no significant neurological or psychological problems who after suffering a GP infarct was subsequently found to have substantial cognitive problems and micrographia. Formal neuropsychological testing was not possible due to lack of patient follow-up. Conclusions and Relevance. Despite the conflicting literature on the association of GP lesions and cognitive deficits, our patient demonstrated significant neuropsychological changes following his stroke. In addition, evidence of micrographia likely adds to the literature on the localization of this finding. Our case thus suggests that neuropsychological testing may be beneficial after GP strokes.

Cdk9 phosphorylates Pirh2 protein and prevents degradation of p53 protein.

Several reports have pointed to the negative involvement of p53 in transcriptional regulation of the human immunodeficiency virus type 1 long-terminal repeat (HIV-1 LTR). We recently demonstrated that through their physical interaction, cdk9 phosphorylates p53 on Ser-392, leading to p53 stability and accumulation. As a result, p53 stalled transcriptional elongation of the HIV-1 LTR and significantly reduced HIV-1 replication in primary microglia and astrocytes. Therefore, we sought to identify the mechanisms used by cdk9 to allow this p53 function. Using western blot analysis, we found that cdk9 promotes inhibition and phosphorylation of Mdm2 on Ser-395, thus preventing degradation of p53, a protein that is directly involved in promoting p53 ubiquitination. On the other hand, we showed that cdk9 phosphorylates Pirh2 on Ser-211 and Thr-217 residues through their physical interaction. Phosphorylation of Pirh2 renders it inactive and may contribute to p53-inhibition of transcriptional elongation of the HIV-1 LTR. Hence, we suggest that phosphorylation of Pirh2 may be a novel target for the inhibition of HIV-1 gene expression.

Role of p53 in neurodegenerative diseases.

BACKGROUND: p53 plays an important role in many areas of cellular physiology and biology, ranging from cellular development and differentiation to cell cycle arrest and apoptosis. Many of its functions are attributed to its role in assuring proper cellular division. However, since the establishment of its role in cell cycle arrest, damage repair, and apoptosis (thus also establishing its importance in cancer development), numerous reports have demonstrated additional functions of p53 in various cells. In particular, p53 appears to have important functions as it relates to neurodegeneration and synaptic plasticity. OBJECTIVE: In this review, we will address p53 functions as it relates to various neurodegenerative diseases, mainly its implications in the development of HIV-associated neurocognitive disorders. CONCLUSION: p53 plays a pivotal role in the development of neurodegenerative diseases through its interaction with cellular factors, viral factors, and/or small RNAs that have the ability to promote the development of these diseases. Hence, inhibition of p53 may present an ideal target to restore neuronal functions.

HIV-1 Tat protein promotes neuronal dysfunction through disruption of microRNAs.

Over the last decade, small noncoding RNA molecules such as microRNAs (miRNAs) have emerged as critical regulators in the expression and function of eukaryotic genomes. It has been suggested that viral infections and neurological disease outcome may also be shaped by the influence of small RNAs. This has prompted us to suggest that HIV infection alters the endogenous miRNA expression patterns, thereby contributing to neuronal deregulation and AIDS dementia. Therefore, using primary cultures and neuronal cell lines, we examined the impact of a viral protein (HIV-1 Tat) on the expression of miRNAs due to its characteristic features such as release from the infected cells and taken up by noninfected cells. Using microRNA array assay, we demonstrated that Tat deregulates the levels of several miRNAs. Interestingly, miR-34a was among the most highly induced miRNAs in Tat-treated neurons. Tat also decreases the levels of miR-34a target genes such as CREB protein as shown by real time PCR. The effect of Tat was neutralized in the presence of anti-miR-34a. Using in situ hybridization assay, we found that the levels of miR-34a increase in Tat transgenic mice when compared with the parental mice. Therefore, we conclude that deregulation of neuronal functions by HIV-1 Tat protein is miRNA-dependent.

Isoform-specific regulation by N(G),N(G)-dimethylarginine dimethylaminohydrolase of rat serum asymmetric dimethylarginine and vascular endothelium-derived relaxing factor/NO.

Asymmetric dimethylarginine (ADMA), which inhibits NO synthase, is inactivated by N(G),N(G)-dimethylarginine dimethylaminohydrolase (DDAH). We tested whether DDAH-1 or -2 regulates serum ADMA (S(ADMA)) and/or endothelium-derived relaxing factor (EDRF)/NO. Small inhibitory (si)RNAs targeting DDAH-1 or -2, or an siRNA control were given intravenously to rats. After 72 hours, EDRF/NO was assessed from acetylcholine-induced, NO synthase-dependent relaxation and 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate for NO activity in isolated mesenteric resistance vessels (MRVs). Expression of mRNA for DDAH-1 versus -2 was 2- and 7-fold higher in the kidney cortex and liver, respectively, whereas expression of DDAH-2 versus -1 was 5-fold higher in MRVs. The proteins and mRNAs for DDAH-1 or -2 were reduced selectively by 35% to 85% in the kidney cortex, liver, and MRVs 72 hours following the corresponding siRNA. S(ADMA) was increased only after siDDAH-1 (266+/-25 versus 342+/-39 [mean+/-SD] nmol x L(-1); P<0.005), whereas EDRF/NO responses and NO activity were not changed consistently by siDDAH-1 but were greatly reduced after siDDAH-2. Mean arterial pressure was not changed significantly by any siRNA. In conclusion, S(ADMA) is regulated by DDAH-1, which is expressed at sites of ADMA metabolism in the kidney cortex and liver, whereas EDRF/NO is regulated primarily by DDAH-2, which is expressed strongly in blood vessels. This implies specific functions of DDAH isoforms.

Role of extracellular superoxide dismutase in the mouse angiotensin slow pressor response.

Low rates of angiotensin II (Ang II) infusion raise blood pressure, renal vascular resistance (RVR), NADPH oxidase activity, and superoxide. We tested the hypothesis that these effects are ameliorated by extracellular superoxide dismutase (EC-SOD). EC-SOD knockout (-/-) and wild type (+/+) mice were equipped with blood pressure telemeters and infused subcutaneously with Ang II (400 ng/kg per minute) or vehicle for 2 weeks. During vehicle infusion, EC-SOD -/- mice had significantly (P<0.05) higher MAP (+/+: 107+/-3 mm Hg versus -/-: 114+/-2 mm Hg; n=11 to 14), RVR, lipid peroxidation, renal cortical p22(phox) expression, and NADPH oxidase activity. Ang II infusion in EC-SOD +/+ mice significantly (P<0.05) increased MAP, RVR, p22(phox), NADPH oxidase activity, and lipid peroxidation. Ang II reduced SOD activity in plasma, aorta, and kidney accompanied by reduced renal EC-SOD expression. During Ang II infusion, both groups had similar values for MAP (+/+ Ang II: 125+/-3 versus -/- Ang II: 124+/-3 mmHg; P value not significant), RVR, NADPH oxidase activity, and lipid peroxidation. SOD activity in the kidneys of Ang II-infused mice was paradoxically higher in EC-SOD -/- mice (+/+: 8.8+/-1.2 U/mg protein(-1) versus -/-: 13.7+/-1.6 U/mg protein(-1); P<0.05) accompanied by a significant upregulation of mRNA and protein for Cu/Zn-SOD. In conclusion, EC-SOD protects normal mice against oxidative stress by attenuating renal p22(phox) expression, NADPH oxidase activation, and the accompanying renal vasoconstriction and hypertension. However, during an Ang II slow pressor response, renal EC-SOD expression is reduced and, in its absence, renal Cu/Zn-SOD is upregulated and may prevent excessive Ang II-induced renal oxidative stress, renal vasoconstriction, and hypertension.

RNA silencing in vivo reveals role of p22phox in rat angiotensin slow pressor response.

The angiotensin II (Ang II) slow-pressor response entails an increase in mean arterial pressure and reactive oxygen species. We used double-stranded interfering RNAs (siRNAs) in Sprague Dawley rats in vivo to test the hypothesis that an increase in the p22phox component of NADPH oxidase is required for this response. Reactive oxygen species were assessed from excretion of 8-isoprostane prostaglandin F2alpha and blood pressure by telemetry. Two siRNA sequences to p22phox (sip22phox) reduced mRNA >85% in cultured vascular smooth muscle cells. Rats received rapid (10 second) IV injections (50 to 100 microg) of 1 of 2 different sip22phox, control siRNA, or vehicle (TransIt in saline) during 14 day SC infusions of Ang II (200 ng.kg(-1).min(-1)) or sham infusions. In both groups, sip22phox, relative to control siRNA, led to significant (P<0.001; approximately 50%) reductions in expression of p22phox mRNA and protein and of NADPH oxidase activity in the kidney cortex. In Ang II-infused rats, sip22phox decreased protein expression for Nox-1, -2, and -4 but increased p47phox. Three days after sip22phox, conscious rats infused with Ang II had a reduced excretion of 8-isoprostane (10+/-1 versus 19+/-2 pg.24 h(-1); P<0.01) and a reduced mean arterial pressure (142+/-5 versus 168+/-4 mm Hg; P<0.005). An increase in p22phox is required for increased renal NADPH oxidase activity, expression of Nox proteins and oxidative stress, and contributes < or =50% to hypertension during an Ang II slow-pressor response.

Expression and cellular localization of classic NADPH oxidase subunits in the spontaneously hypertensive rat kidney.

Phagocytes generate superoxide anion (O(2)(-)) by a classic, 5-component NADPH oxidase. O(2)(-) contributes to hypertension in spontaneously hypertensive rats (SHR). Therefore, we tested the hypothesis that NADPH oxidase expression is enhanced in the SHR kidney. We also analyzed the localization of NADPH oxidase components in SHR kidney. Renal NADPH oxidase was quantified by reverse transcription-polymerase chain reaction and Western blotting and was localized in SHR and Wistar Kyoto rat (WKY) kidney by immunohistochemistry. The mRNA for 5 subunits of phagocyte NADPH oxidase, and also for MOX1 and RENOX (NOX4), was detected in adult rat kidney. Kidneys of adult (10 weeks old) SHR had a significantly (P<0.01) greater mRNA for p47phox (SHR 0.81 +/- 0.05 versus WKY 0.37 +/- 0.01, arbitrary unit), which was confirmed by Western blotting (SHR 0.58 +/- 0.04 versus WKY 0.42 +/- 0.04, arbitrary unit; P<0.05) and by immunohistochemistry. This higher p47phox protein expression was also detected in young prehypertensive SHR (SHR 0.61 +/- 0.05 versus WKY 0.39 +/- 0.04, arbitrary unit; P<0.01). The 10-week-old SHR contained more modest but significantly (P<0.05) greater protein for p67phox (SHR 0.54 +/- 0.02 versus WKY 0.46 +/- 0.02). Immunostaining localized p47phox, p67phox, and p22phox in vasculature, macula densa, distal convoluted tubule, cortical collecting duct, and outer and inner medullary collecting ducts. The kidney of SHR expresses genes for all the main components of phagocyte NADPH oxidase, RENOX, and MOX1. There is a prominent increase in the SHR kidney of the mRNA, and protein expression of p47phox in the vasculature, macula densa, and distal nephron, which precedes development of hypertension.