FETAL ASCITES A Report of 3 Autopsy Cases

Three rare autopsy cases of fetal ascites were presented and the etiology of each case was described. Case 1 was a male neonate, delivered by cesarean section at 32 weeks’gestation, and died of respiratory failure. The abdomen was remarkably distended with 1020 ml of ascites. The etiology of Case 1 remained unknown even after macroscopic and microscopic examinations. We considered this as “idiopathic” fetal ascites. Case 2 was a female neonate, delivered at 31 weeks’gestation, with marked abdominal distension and cyanosis. Autopsy revealed 435 ml of ascites, and she was considered to have had “polysplenia syndrome” with cardiovascular malformations. Intrauterine heart failure due to cardiac anomalies was thought to be the cause of this ascites. In case 3 embryotomy was carried out under the diagnosis of fetal ascites by ultrasound examination at 22 weeks’gestation. An urachal cyst connected to the dilated urinary bladder and deficiency of musculature of the abdominal wall composed of loose connective tissue with calcification were observed. The abdominal wall was ruptured and 1,960 ml of ascites was measured. Polycystic kidney with renal dysplasia was also found. Case 3 showed “Prune-Berry syndrome” and fetal ascites may have arisen from these anomalies.     

Picoliter‐Droplet Digital Polymerase Chain Reaction‐Based Analysis of Cell‐Free Plasma DNA to Assess EGFR Mutations in Lung Adenocarcinoma That Confer Resistance to Tyrosine‐Kinase Inhibitors

Purpose.

The objective of this study was to evaluate the utility of analyzing cell-free plasma DNA (cfDNA) by picoliter-droplet digital polymerase chain reaction (ddPCR) to detect EGFR mutations that confer resistance to tyrosine-kinase inhibitors (TKIs) used for treatment of lung adenocarcinoma (LADC).

Experimental design.

Thirty-five LADC patients who received epidermal growth factor receptor (EGFR)-TKI therapy, including ten who received tumor rebiopsy after development of resistance, were subjected to picoliter-ddPCR-cfDNA analysis to determine the fraction of cfDNA with TKI-sensitive (L858R and inflame exon 19 deletions) and -resistant (i.e., T790M) mutations, as well as their concordance with mutation status in rebiopsied tumor tissues.

Results.

cfDNA samples from 15 (94%) of 16 patients who acquired resistance were positive for TKI-sensitive mutations. Also, 7 (44%) were positive for the T790M mutation, with fractions of T790M (+) cfDNA ranging from 7.4% to 97%. T790M positivity in cfDNA was consistent in eight of ten patients for whom rebiopsied tumor tissues were analyzed, whereas the remaining cases were negative in cfDNA and positive in rebiopsied tumors. Prior to EGFR-TKI therapy, cfDNAs from 9 (38%) and 0 of 24 patients were positive for TKI-sensitive and T790M mutations, respectively. Next-generation sequencing of cfDNA from one patient who exhibited innate resistance to TKI despite a high fraction of TKI-sensitive mutations and the absence of the T790M mutation in his cfDNA revealed the presence of the L747P mutation, a known driver of TKI resistance.

Conclusion.

Picoliter-ddPCR examination of cfDNA, supported by next-generation sequencing analysis, enables noninvasive assessment of EGFR mutations that confer resistance to TKIs.

Implications for Practice:

Noninvasive monitoring of the predominance of tumors harboring the secondary T790M mutation in the activating mutation in EGFR gene is necessary for precise and effective treatment of lung adenocarcinoma. Because cells harboring the T790M mutation are resistant to epidermal growth factor receptor-tyrosine-kinase inhibitors (TKIs), the predominance of tumor cells harboring the T790M mutations influences the choice of whether to use conventional or next-generation TKIs. Digital polymerase chain reaction-based examination of cfDNA is a promising method; however, its feasibility, including its consistency with examination of rebiopsied tumor tissue, has not been fully proven. Here, picoliter-droplet digital polymerase chain reaction technology is presented as a candidate method for testing cfDNA and assessing the predominance of T790M-mutant tumors.     

Identification of anti‐cancer chemical compounds using Xenopus embryos

Cancer tissues have biological characteristics similar to those observed in embryos during development. Many types of cancer cells acquire pro‐invasive ability through epithelial–mesenchymal transition (EMT). Similar processes (gastrulation and migration of cranial neural crest cells [CNCC]) are observed in the early stages of embryonic development in Xenopus during which cells that originate from epithelial sheets through EMT migrate to their final destinations. The present study examined Xenopus embryonic tissues to identify anti‐cancer compounds that prevent cancer invasion. From the initial test of known anti‐cancer drugs, AMD3100 (an inhibitor of CXCR4) and paclitaxel (a cytoskeletal drug targeting microtubules) effectively prevented migration during gastrulation or CNCC development. Blind‐screening of 100 synthesized chemical compounds was performed, and nine candidates that inhibited migration of these embryonic tissues without embryonic lethality were selected. Of these, C‐157 (an analog of podophyllotoxin) and D‐572 (which is an indole alkaroid) prevented cancer cell invasion through disruption of interphase microtubules. In addition, these compounds affected progression of mitotic phase and induced apoptosis of SAS oral cancer cells. SAS tumors were reduced in size after intratumoral injection of C‐157, and peritoneal dissemination of melanoma cells and intracranial invasion of glioma cells were inhibited by C‐157 and D‐572. When the other analogues of these chemicals were compared, those with subtle effect on embryos were not tumor suppressive. These results suggest that a novel chemical‐screening approach based on Xenopus embryos is an effective method for isolating anti‐cancer drugs and, in particular, targeting cancer cell invasion and proliferation.     

Monocyte infiltration into obese and fibrilized tissues is regulated by PILRα

Paired immunoglobulin‐like type 2 receptor α (PILRα) is an inhibitory receptor that is mainly expressed on myeloid cells, and negatively regulates neutrophil infiltration during inflammation. However, PILRα role on monocyte has not been described. Under both steady‐state and inflammatory conditions, monocytes migrate into tissues and differentiate into macrophages. Macrophages in adipose and liver tissues play important roles in tissue homeostasis and pathogenesis of metabolic diseases. Here, we found that PILRα controls monocyte mobility through regulating integrin signaling and inhibiting CD99–CD99 binding. Moreover, we found that Pilra^?/? mice developed obesity and hepatomegaly with fibrosis, and the numbers of macrophages in adipose and liver tissues are significantly increased in Pilra^?/? mice. These data suggest that immune inhibitory receptor, PILRα, plays an important role in the prevention of obesity and liver fibrosis.     

REM Sleep Components Predict the Response to Initial Treatment of Infantile Spasms

PURPOSE: The phasic inhibition index (PII) is the rate of the simultaneous occurrence of rapid eye movement bursts (RBs) and phasic chin muscle activity (PCMA) during rapid eye movement sleep (REMS). PII is low insofar as physiologically occurring REM-related phasic inhibition acts on chin muscles. Previously we found that PII was significantly higher in patients with infantile spasms (ISs) who had a recurrence of convulsions than in patients with ISs who exhibited no recurrence. We aimed to predict the response of patients with ISs to conventional anticonvulsants (AEDs) by means of REMS components including PII, expecting to facilitate avoidance of potentially hazardous hormonal therapy. METHODS: REMS, recorded before the beginning of any medication, was retrospectively examined in 15 patients with ISs. The patients were classified into two groups according to the response to initial treatment with conventional AEDs. Conventional AEDs were enough to control the spasms in six good responders (GRs), whereas further hormonal therapy was required in nine poor responders (PRs) to control the spasms. RESULTS: The amount of REMS was significantly lower in patients with ISs than in controls. GRs had less REMS than did PRs, although no significant difference was observed. Although the frequencies of RB and PCMA showed no significant differences among GRs, PRs, and controls, the average PII value in PRs (12.6+/-3.4; mean+/-SD) was significantly (p < 0.001) higher than that in GRs (6.1+/-1.7). CONCLUSIONS: PII is a useful parameter for differentiating GRs from PRs.     

Diagnosis and assessment of preclinical and clinical autoimmune encephalomyelitis using peripheral blood lymphocyte TCR

In organ-specific autoimmune diseases, T cells involved in the disease development bear a particular type of TCR and infiltrate the target organ predominantly. However, it is difficult to identify disease-inducing T cells in peripheral blood lymphocytes (PBL) because such T cells are very few in number in a large pool of unrelated T cells. In the present study, we demonstrate that CDR3 spectratyping can identify experimental autoimmune encephalomyelitis (EAE)-specific patterns (oligoclonal expansion of Vβ8.2 with the shortest CDR3) in PBL at the preclinical and clinical stages of acute EAE. Analysis of nucleotide and predicted amino acid sequences of Vβ8.2 CDR3 of spectratype-derived clones revealed that CASSDSSYEQYFGPG, which is one of the representative sequences of encephalitogenic T cell clones, constituted the predominant population in both PBL and spinal cord T cells. In chronic relapsing EAE, the EAE-specific spectratype pattern in PBL was observed during the 1st and 2nd attacks, but not at the remission and full recovery stage. These findings indicate that the spectratyping pattern in PBL reflects the disease activity of acute and chronic relapsing EAE. Thus, CDR3 spectratyping using PBL can be used for diagnosis and assessment of T cell-mediated autoimmune diseases and is applicable to human autoimmune diseases.