Gut microbiota development of preterm infants hospitalised in intensive care units.

Gut microbiome development affects infant health and postnatal physiology. The gut microbe assemblages of preterm infants have been reported to be different from that of healthy term infants. However, the patterns of ecosystem development and inter-individual differences remain poorly understood. We investigated hospitalised preterm infant gut microbiota development using 16S rRNA gene amplicons and the metabolic profiles of 268 stool samples obtained from 17 intensive care and 42 term infants to elucidate the dynamics and equilibria of the developing microbiota. Infant gut microbiota were predominated by Gram-positive cocci, Enterobacteriaceae or Bifidobacteriaceae, which showed sequential transitions to Bifidobacteriaceae-dominated microbiota. In neonatal intensive care unit preterm infants (NICU preterm infants), Staphylococcaceae abundance was higher immediately after birth than in healthy term infants, and Bifidobacteriaceae colonisation tended to be delayed. No specific NICU-cared infant enterotype-like cluster was observed, suggesting that the constrained environment only affected the pace of transition, but not infant gut microbiota equilibrium. Moreover, infants with Bifidobacteriaceae-dominated microbiota showed higher acetate concentrations and lower pH, which have been associated with host health. Our data provides an in-depth understanding of gut microbiota development in NICU preterm infants and complements earlier studies. Understanding the patterns and inter-individual differences of the preterm infant gut ecosystem is the first step towards controlling the risk of diseases in premature infants by targeting intestinal microbiota.

Periodontal healing after replantation of intentionally rotated teeth with healthy and denuded root surfaces.

The purpose of the study was to evaluate periodontal healing after replantation of intentionally rotated teeth with healthy and denuded root surfaces. Ten teeth with hopeless prognoses because of extensive alveolar bone loss and deep pockets extending to the apexes only on one or two surfaces, but with a healthy periodontal apparatus on the other surfaces, were selected. A mucogingival flap was elevated and the teeth were extracted carefully so as not to damage the healthy remnant of the periodontal membrane remaining on the root surfaces. Thorough extra-oral debridement was performed on the contaminated root surface and the remnant was carefully conserved. Retrofilling was also done to eliminate an endodontic cause of attachment loss. The teeth were horizontally rotated and then replanted so that the healthy root would face the connective tissue at the initially periodontally involved sites, and the root planed surfaces, which had been periodontally involved, would face the surfaces of the alveolar sockets at initially healthy sites. The teeth were splinted with adjacent teeth. Clinical parameters and radiographic examination were performed pre-operatively, and at 3 or 6 months, 1, 2, and 3 years postoperatively. One tooth was extracted 1.5 years postoperatively due to reduced support and the treatment strategy of the patient. The other nine teeth were well maintained without any discomfort for the whole maintenance period of 3 years. In all teeth, areas that previously had no pocketing but were now against denuded root surfaces maintained the previous pocket depth readings. In areas where a deep pocket was present but now had a root surface with healthy periodontium, a distinct decrease of mean pocket depth was observed at the first reexamination 3 months postoperatively. The mean radiographic alveolar bone level increased from 0.3% to 45.3% in 2 years and was thereafter maintained for the entire observation period. Loss of periodontal space and possible root resorption were observed in only one case without other ankylotic symptoms. These results suggest that the healing without ankylosis of an extensive denuded root surface may occur by mechanisms other than the maintenance of a viable periodontal ligament on the root surface.

Doxycycline reduction of F-actin content of human neutrophils and fibroblasts.

We and others have shown that tetracyclines inhibit leukocyte adherence, migration, and phagocytosis, functions presumed to involve actin microfilament metabolism. In this study we investigated the influence of a tetracycline (doxycycline, Dc) on neutrophil and fibroblast actin metabolism. Human neutrophils and fibroblasts were pretreated with Dc or cytochalasin B (cB), stimulated with either the chemotactic peptide FMLP or phorbol myristate acetate (PMA), and the changes in phalloidin conjugate, associated F-actin were followed microscopically or quantified fluorometrically. Doxycycline suppressed, in a dose-related manner, the rise in F-actin content of neutrophils that normally follows their activation with either FMLP or PMA. Cytochalasin B had a similar affect on actin microfilament synthesis. Incubation of fibroblasts in Dc led to a loss of actin microfilaments and caused flattened adherent cells to round up and detach. Both Dc and cB also inhibited Con A-induced acceptor capping on neutrophils, a phenomenon known to be dependent on the presence of intact actin microfilaments. The data show that both Dc and cB influence actin metabolism and suggest they do so by differing mechanisms.

The influence of doxycycline of the attachment of fibroblasts to gelatin-coated surfaces and its cytotoxicity.

Because of their antimicrobial and anti-inflammatory properties, tetracyclines have been used systemically and locally in the treatment of periodontal disease. This study was done to evaluate the influence of doxycycline (Dc), a tetracycline, on fibroblast adherence to a protein coated surface and its cytotoxicity. Periodontal ligament derived fibroblasts (PLDFs) were either: 1) preincubated with Dc (0 to 100 micrograms/ml) and then allowed to adhere to a gelatin-coated surface or 2) adherence was first established and then Dc added to the system. After an appropriate time the number of PLDFs adherent, released, or lysed was estimated by a lactic dehydrogenase (LDH) assay. Preincubation of PLDFs in 25 micrograms Dc/ml or higher concentrations significantly (P < 0.01) reduced the number of adherent cells. Fifty micrograms Dc/ml and higher doses significantly (P < 0.01) increased PLDFs cytotoxicity as measured by LDH release. The same trend was noted if PLDFs were allowed to adhere and then subjected to the drug. Microscopic observation of fluorescein diacetate/propidium iodide-stained cells showed that attached-spread cells pulled in, rounded up, and detached in proportion to the increased dose of Dc and the percentage of red-stained cytotoxic cells rose in a similar manner. The data suggested that Dc can be cytotoxic and may inhibit PLDFs adherence and spread along a substratum.

Factors relating to coagulation, fibrinolysis and hepatic damage after liver resection.

A survey of the blood of twenty-two patients who had undergone hepatic resection was performed. Serum levels of alpha-2 plasmin inhibitor-plasmin complex initially decreased from 1.58 +/- 0.31 microgram/ml on the preoperative day (PREOP), to 0.92 +/- 0.14 mu/ml on the first postoperative day (POD 1), and then increased to 3.13 +/- 0.92 micrograms/ml on the seventh postoperative day (POD 7) (mean +/- SE)). Thrombin-anti-thrombin III complex (14.2 +/- 4.3 ng/ml on PREOP and 26.0 +/- 4.1 ng/ml on POD 7 (mean +/- SE)) and D-dimer (335 +/- 96 ng/ml on PREOP and 1859 +/- 258 ng/ml on POD 7 (mean +/- SE)) increased in the early postoperative stage. The level of 6-keto-prostaglandin F1 alpha increased after the operations (from 13.2 +/- 1.8 pg/ml on PREOP to 37.8 +/- 12.8 pg/ml on POD 7 (mean +/- SE)). The level of thromboxane B-2 decreased at first, and then gradually increased and returned to its preoperative level on POD 7 (144.7 +/- 43.8 pg/ml on PREOP, 57.6 +/- 27.5 pg/ml on POD 1 and 152.5 +/- 58.4 pg/ml on POD 7 (mean +/- SE)). Superoxide dismutase activity increased at first, and then gradually decreased, postoperatively (2.8 +/- 0.5 NU/ml on PREOP, 4.8 +/- 0.8 NU/ml on POD 1 and 2.6 +/- 0.3 NU/ml on POD 7 (mean +/- SE)). That is, biodefensive reactions which protect patients against the shift to disseminated intravascular coagulation (DIC) were inferred with by the increase in antiplatelet aggregation, despite the activation of coagulation and fibrinolytic mechanisms after hepatic resection.

Comparison of doxycycline and a chemically modified tetracycline inhibition of leukocyte functions.

Tetracyclines (Tc's) have the ability to inhibit neutrophil (PMN) functions which may be of value in the treatment of neutrophil-driven pathologic processes. However, long term use of Tc's frequently lead to emergence of antibiotic resistant strains of bacteria. A chemically modified analogue of Tc, 4DTc, having minimal antibiotic activity was compared to Dc, a member of the Tc family, for its ability to inhibit neutrophil functions. 4DTc was significantly less effective at inhibiting PMN superoxide synthesis, PMA-induced degranulation and PMN-mediated RBC lysis than was Dc. 4DTc and Dc were equally as effective inhibiting monocyte, but not PMN, adherence to gelatin-coated surfaces. When incubated in media containing varying 4DTc or Dc concentrations, Dc accumulated in RBC's and PMN's at levels 3 times greater than that found for similar media levels of 4DTc. The data suggest that the lesser ability of 4DTc to inhibit several PMN functions as compared to Dc may be related to its reduced intracellular accumulation. It also suggest that Tc inhibition of monocyte adherence may be more influenced by extracellular than intracellular processes.

[Superior mesenteric arterial infusion chemotherapy in patient with both peritoneal carcinomatosis and liver metastasis].

Experimental study of superior mesenteric arterial infusion chemotherapy was done to evaluate the effect of this therapy on metastatic liver tumor. This treatment has been tried clinically for ileus caused by peritoneal carcinomatosis with good result. Moreover, the infused drug is expected to reach the liver via portal route, just like intraportal administration. This study disclosed that superior mesenteric arterial infusion made for a sufficiently high 5-FU concentration of portal blood but the concentration in the metastatic liver tumor was lower when compared with intravenous infusion. However, because the 5-FU concentration in blood of the aorta was lower than in the intravenous administration group, our method may decrease the unfavorable side effects of anti-cancer drugs.

The influence of divalent cations and doxycycline on iodoacetamide-inhibitable leukocyte adherence.

Leukocyte adherence, an event critical to host defense, is reported to be dependent upon divalent cations. To test whether leukocyte binding is more influenced by the availability of Ca2+ or Mg2+, neutrophils and mononuclear leukocyte adherence to surfaces coated with differing proteins, in medium containing varying concentrations of Ca2+ and/or Mg2+ and induced with either FMLP or PMA were assessed. To account for nonspecific leukocyte-substrata interactions the OD's of iodoacetamide-inhibited adherent cell systems were subtracted from the OD values of adherent cells of noninhibited systems. Values presented were derived from OD's of iodoacetamide-inhibitable leukocyte binding. PMA was a much more potent inducer of leukocyte adherence than was FMLP, stimulating all available neutrophils and monocytes to adhere. In contrast, FMLP induced adherence by roughly 1/4 of the available neutrophils and few, if any, monocytes. Roughly the same binding pattern was noted whether surfaces were coated with albumin, fibrinogen, fibronectin, gelatin or serum. EDTA but not EGTA significantly suppressed leukocyte binding suggesting Mg2+ was more involved in binding than was Ca2+. Little leukocyte adherence to a substrata occurred in the absence of divalent cations, while enrichment of the medium with Mg2+ was more influential on cell binding than was enrichment with Ca2+. Interestingly, doxycycline, a member of the tetracycline family of drugs which has been reported to inhibit Mg(2+)-dependent neutrophil functions, had a slight inhibitory effect on neutrophil adherence at low drug concentrations, while it enhanced binding at higher doxycycline concentrations. In contrast, the tetracycline inhibited monocyte adherence in a dose-related manner.

[Clinical study of drug accumulation in gastric cancer after preoperative intra-arterial EAP II injection therapy].

Preoperative intra-arterial injection therapy using etoposide, epirubicin and carboplatin (EAP II) was done for patients with resectable advanced gastric cancer. Twenty-six patients (14 males and 12 females) were treated. The concentrations of adriamycin (ADM) and platinum (Pt) were measured in cancer tissue, normal mucosa and regional lymph-nodes which were obtained operatively and in sera just before operation. But the concentration of etoposide was not measured, because when we used preoperative intra-arterial injection therapy using etoposide, epirubicin and cisplatin (EAP I), the mean concentration of etoposide was less than the detectable limit in all tissues and in sera. There were no significant differences among the mean concentrations of ADM in all tissues and in sera. And the mean concentration of ADM in cancer tissue was not higher than that of intra-arterial EAP I injection therapy. The mean concentration of platinum in sera was significantly lower than in cancer tissues, normal mucosa and lymph-nodes. And the mean concentrations of platinum in cancer tissues and lymph-nodes were higher than those of intra-arterial CDDP or EAP I injection therapy. It was concluded that preoperative intra-arterial EAP II injection therapy may be an effective method to improve the usefulness of preoperative intra-arterial injection therapy for gastric cancer.

[Superior mesenteric arterial infusion chemotherapy against intestinal obstruction caused by peritoneal carcinomatosis].

For 4 patients with peritoneal carcinomatosis who had severe abdominal pain and vomiting, intra-arterial infusion chemotherapy, via the superior mesenteric artery was performed. After the treatment, all patients were free of their symptoms and began to eat again. Severe complications such as superior mesenteric arterial thrombosis did not occur. It is concluded that our treatment is clinically useful because the quality of life of these patients at the end stage was improved.

[Clinical study of the drug delivery in metastatic liver cancer using bromodeoxyuridine].

As the model of an anti-cancer agent acting at DNA synthesizing phase, BrdU was infused for metastatic liver cancers into the portal vein and/or the hepatic artery. After administration, the liver tumors were resected, and immunohistochemical staining was performed using anti-BrdU monoclonal antibody. Two of 6 portal group tumors were stained. All 4 arterial and 3 arterio-portal group tumors were stained. Therefore, it was proved that metastatic liver tumors had both arterial and portal blood supply. But areas to which BrdU was delivered were only peripheral, and the inmost area was 2 mm from the tumor surface. After the intra-arterial infusion of BrdU suspended in lipiodol, the delivery of BrdU was highly enhanced and 4 of 6 tumors were stained even to the deepest areas. However, one tumor of which the major part was necrotic and one tumor which produced mutin were not stained at all. It was interesting that after intra-arterial administration of BrdU suspended in lipiodol, BrdU was also found in the cytoplasm of normal liver cells.

[Clinical study of platinum accumulation in gastric cancer after preoperative intra-arterial injection of cisplatin].

In order to deliver a high concentration of CDDP in tumor tissue, we attempted to infuse CDDP intra-arterially for preoperative patients with resectable gastric cancer. Thirty-two patients (21 males and 11 females) were treated. The concentration of platinum was measured in cancer tissue, normal mucosa, lymph nodes without metastasis at the greater curvature. These were gathered operatively and in serum just before operation. The serum concentration of free-CDDP was also measured. Student's-t test was performed with the data. After intra-arterial injection of 60 mg CDDP, the mean concentration of platinum in cancer tissue was 1.11 +/- 0.45 microgram/g and after intravenous injection of 40 mg CDDP that was 0.30 +/- 0.11 microgram/g. After intra-arterial injection of 40 mg CDDP, the mean concentration of platinum in cancer tissue was 0.64 +/- 0.12 microgram/g, which was significantly higher than in normal gastric mucosa (0.27 +/- 0.06 micrograms/g) or serum (0.37 +/- 0.10 micrograms/ml) (p less than 0.025). The mean concentration of platinum in cancer tissue was higher than in the lymph nodes (0.45 +/- 0.10 micrograms/g), but there was no significant difference. In sera the concentrations of free-CDDP were all under the detectable limit. It was concluded that intra-arterial injection therapy of CDDP was an effective method to maintain a high concentration of CDDP in gastric cancer tissue.

[A case of breast carcinoma, possibly the result of malignant transformation of an intraductal papilloma in a 26-year period].

A 74-year old female presented a giant tumor and serosanguineous nipple discharge from the left breast. The tumor was first recognized 26 years ago, and untreated. The resected breast contained several cysts and some small white nodules. Some of the cysts contained intracystic tumors. Histologically, the tumor consisted of a mixture of papillotubular carcinoma and intraductal papilloma. This case seems to represent a malignant change in intraductal papilloma from its long clinical course and by the microscopic findings. Review of this case seems to warrant clinical attention to the possibility that benign intraductal papilloma may transform itself into malignancy after many years.