CDK1 Is a Synthetic Lethal Target for KRAS Mutant Tumours

Activating KRAS mutations are found in approximately 20% of human cancers but no RAS-directed therapies are currently available. Here we describe a novel, robust, KRAS synthetic lethal interaction with the cyclin dependent kinase, CDK1. This was discovered using parallel siRNA screens in KRAS mutant and wild type colorectal isogenic tumour cells and subsequently validated in a genetically diverse panel of 26 colorectal and pancreatic tumour cell models. This established that the KRAS/CDK1 synthetic lethality applies in tumour cells with either amino acid position 12 (p.G12V, pG12D, p.G12S) or amino acid position 13 (p.G13D) KRAS mutations and can also be replicated in vivo in a xenograft model using a small molecule CDK1 inhibitor. Mechanistically, CDK1 inhibition caused a reduction in the S-phase fraction of KRAS mutant cells, an effect also characterised by modulation of Rb, a master control of the G₁/S checkpoint. Taken together, these observations suggest that the KRAS/CDK1 interaction is a robust synthetic lethal effect worthy of further investigation.     

Receptor for Advanced Glycation End Products (RAGE) Serves a Protective Role during Klebsiella pneumoniae - Induced Pneumonia

Klebsiella species is the second most commonly isolated gram-negative organism in sepsis and a frequent causative pathogen in pneumonia. The receptor for advanced glycation end products (RAGE) is expressed on different cell types and plays a key role in diverse inflammatory responses. We here aimed to investigate the role of RAGE in the host response to Klebsiella (K.) pneumoniae pneumonia and intransally inoculated rage gene deficient (RAGE^-/-) and normal wild-type (Wt) mice with K. pneumoniae. Klebsiella pneumonia resulted in an increased pulmonary expression of RAGE. Furthermore, the high-affinity RAGE ligand high mobility group box-1 was upregulated during K. pneumoniae pneumonia. RAGE deficiency impaired host defense as reflected by a worsened survival, increased bacterial outgrowth and dissemination in RAGE^-/- mice. RAGE^-/- neutrophils showed a diminished phagocytosing capacity of live K. pneumoniae in vitro. Relative to Wt mice, RAGE^-/- mice demonstrated similar lung inflammation, and slightly elevated—if any—cytokine and chemokine levels and unchanged hepatocellular injury. In addition, RAGE^-/- mice displayed an unaltered response to intranasally instilled Klebsiella lipopolysaccharide (LPS) with respect to pulmonary cell recruitment and local release of cytokines and chemokines. These data suggest that (endogenous) RAGE protects against K. pneumoniae pneumonia. Also, they demonstrate that RAGE contributes to an effective antibacterial defense during K. pneumoniae pneumonia, at least partly via its participation in the phagocytic properties of professional granulocytes. Additionally, our results indicate that RAGE is not essential for the induction of a local and systemic inflammatory response to either intact Klebsiella or Klebsiella LPS.     

Some characteristics of pathogenic avian PPLO

Summary Pathogenic avian PPLO,Mycoplasma gallisepticum strains were seen to grow on solid media in two types of colonies. The small ones are typical PPLO, the large ones seem to occupy an intermediate position between PPLO and bacteria. The elements constituting the large colonies must be the so-called Nelson bodies.     

Ketamine Does Not Produce Relief of Neuropathic Pain in Mice Lacking the β-Common Receptor (CD131)

Neuropathic pain (NP) is a debilitating condition associated with traumatic, metabolic, autoimmune and neurological etiologies. Although the triggers for NP are diverse, there are common underlying pathways, including activation of immune cells in the spinal cord and up-regulation of the N-methyl-D-aspartate receptor (NMDAR). Ketamine, a well-known NDMAR antagonist, reduces neuropathic pain in a sustained manner. Recent study has shown that the novel 11-amino acid peptide erythropoietin derivative ARA290 produces a similar, long-lasting relief of NP. Here, we show that both drugs also have similar effects on the expression of mRNA of the NMDAR, as well as that of microglia, astrocytes and chemokine (C-C motif) ligand 2, all-important contributors to the development of NP. Although the effects of ketamine and ARA 290 on NP and its molecular mediators suggest a common mechanism of action, ARA 290 has no affinity for the NMDAR and acts specifically via the innate repair receptor (IRR) involved in tissue protection. We speculated therefore, that the IRR might be critically involved in the action of ketamine on neuropathic pain. To evaluate this, we studied the effects of ketamine and ARA 290 on acute pain, side effects, and allodynia following a spared nerve injury model in mice lacking the β-common receptor (βcR), a structural component of the IRR. Ketamine (50 mg/kg) and ARA 290 (30 µg/kg) produced divergent effects on acute pain: ketamine produced profound antinociception accompanied with psychomotor side effects, but ARA290 did not, in both normal and knock out mice. In contrast, while both drugs were antiallodynic in WT mice, they had no effect on NP in mice lacking the βcR. Together, these results show that an intact IRR is required for the effective treatment of NP with either ketamine or ARA 290, but is not involved in ketamine’s analgesic and side effects.     

Psychophysiological Response Patterns to Affective Film Stimuli

Psychophysiological research on emotion utilizes various physiological response measures to index activation of the defense system. Here we tested 1) whether acoustic startle reflex (ASR), skin conductance response (SCR) and heart rate (HR) elicited by highly arousing stimuli specifically reflect a defensive state and 2) the relation between resting heart rate variability (HRV) and affective responding. In a within-subject design, participants viewed film clips with a positive, negative and neutral content. In contrast to SCR and HR, we show that ASR differentiated between negative, neutral and positive states and can therefore be considered as a reliable index of activation of the defense system. Furthermore, resting HRV was associated with affect-modulated characteristics of ASR, but not with SCR or HR. Interestingly, individuals with low-HRV showed less differentiation in ASR between affective states. We discuss the important value of ASR in psychophysiological research on emotion and speculate on HRV as a potential biological marker for demarcating adaptive from maladaptive responding.     

The role of human innate immune factors in nasal colonization by Staphylococcus aureus

Staphylococcus aureus colonization of the human nares predisposes to sometimes severe auto-infection. To investigate whether genetic polymorphism affects the S. aureus carriage status, sequence variation in alpha-defensin and beta-defensin, and mannose-binding lectin (MBL) genes were determined for a group of volunteers (n=109) with known S. aureus nasal carriage status. DEFA1/3 expression was measured in a subset of the volunteers (n=32). None of the single nucleotide polymorphisms studied could clearly distinguish the (non) carriage groups. S. aureus carriers differed from non-carriers in baseline level of HNP1-3 peptide production (median: 218 versus 89mug/ml, P=0.016). No association between HNP1-3 levels and the individual sequence polymorphisms was documented. The combined copy numbers of DEFA1/A3 genes ranged from 5 to 23 per diploid genome. A linear correlation between combined copy numbers and HNP1-3 peptide concentrations in nasal secretions of non-carriers was noted (r(2)=0.8991). DEFA3 gene was absent in 25% of the individuals. MBL haplotype A was overrepresented in persistent S. aureus carriers (87% vs. 67%; P=0.038). In conclusion, defensin gene polymorphism, both in sequence and in gene copy numbers, does not seem to be involved in S. aureus carriage predisposition. However, MBL haplotypes do so significantly. Baseline HNP1-3 production is more the consequence of S. aureus colonization than a reason for the (non) carrier status.     

Positive Feedbacks in Seagrass Ecosystems: Implications for Success in Conservation and Restoration

Abstract Seagrasses are threatened by human activity in many locations around the world. Their decline is often characterized by sudden ecosystem collapse from a vegetated to a bare state. In the 1930s, such a dramatic event happened in the Dutch Wadden Sea. Before the shift, large seagrass beds (Zostera marina) were present in this area. After the construction of a large dam and an incidence of the “wasting disease” in the early 1930s, these meadows became virtually extinct and never recovered despite restoration attempts. We investigated whether this shift could be explained as a critical transition between alternative stable states, and whether the lack of recovery could be due to the high resilience of the new turbid state. We analyzed the depth distribution of the historical meadows, a long-term dataset of key factors determining turbidity and a minimal model based on these data. Results demonstrate that recovery was impossible because turbidity related to suspended sediment was too high, probably because turbidity was no longer reduced by seagrass itself. Model simulations on the positive feedback suggest indeed the robust occurrence of alternative stable states and a high resilience of the current turbid state. As positive feedbacks are common in seagrasses, our findings may explain both the worldwide observed collapses and the low success rate of restoration attempts of seagrass habitats. Therefore, appreciation of ecosystem resilience may be crucial in seagrass ecosystem management.     

Over expression of IL-10 by macrophages overcomes resistance to murine filariasis

Individuals infected with parasitic helminths are able to tolerate the presence of parasites for considerable time without clinical pathology. Immunosuppressive responses induced by the filarial parasite are considered responsible for this long-lasting relationship, inuring to the benefit of both parasite and host. In order to directly link IL-10 with parasite survival, we infected mice, in which over expression of IL-10 was restricted to macrophages under control of the CD68 promoter (macIL-10tg), with Litomosoides sigmodontis. IL-10 overexpression by macrophages led to increased susceptibility with a significantly higher number of adult worms. Most profound, IL-10 overexpression was sufficient to convert resistant FVB wild-type mice towards a patent phenotype, since microfilariae were exclusively found in macIL-10tg mice. These findings were associated with reduced Th2 cytokine production in macIL-10tg mice. Expression of arginase-1, Ym1 and Fizz1, genes that are found strongly expressed in murine alternatively activated macrophages, were detected in macIL-10tg mice. Thus, IL-10 produced by macrophages with characteristics of alternative activation can overcome resistance and allow full patency in murine filariasis.