Single-cell analysis of human dermal fibroblasts isolated from a single male donor over 35 years.

The aim of this study is to examine the effects of ageing on dermal fibroblast heterogeneity based on samples obtained from the same donor. We used a dermal fibroblast lineage (named ASF-4 cell lines) isolated from the inner side of the upper arm of a healthy male donor over a 35-year period, beginning at 36 years of age. Because clonal analysis of ASF-4 cell lines demonstrated a donor age-dependent loss of proliferative capacity and acquisition of senescent traits at the single-cell level, cultured cells frozen at passage 10 at ages 36 and 72 years were subjected to single-cell RNA sequencing. Transcriptome analysis revealed an increase in senescent fibroblasts and downregulation of genes associated with extracellular matrix remodelling with ageing. In addition, two putative differentiation pathways, with one endpoint consisting of senescent fibroblasts and the other without, were speculated using a pseudo-time analysis. Knockdown of the characteristic gene of the non-senescent fibroblast cluster endpoint, EFEMP2, accelerated cellular senescence. This was also confirmed in two other normal human dermal fibroblast cell lines. The detection of a common cellular senescence-related gene from single-donor analysis is notable. This study provides new insights into the behaviour of dermal fibroblasts during skin ageing.

Three-dimensional structure analysis of melanocytes and keratinocytes in senile lentigo.

Senile lentigo or age spots are hyperpigmented macules of skin that commonly develop following long-term exposure to ultraviolet radiation. This condition is caused by accumulation of large numbers of melanosomes (melanin granules) produced by melanocytes within neighboring keratinocytes. However, there is still no consensus regarding the melanosome transfer mechanism in senile lentigo. To date, most pathohistological studies of skin have been two-dimensional and do not provide detailed data on the complex interactions of the melanocyte-keratinocyte network involved in melanosome transfer. We performed a three-dimensional reconstruction of the epidermal microstructure in senile lentigo using three different microscopic modalities to visualize the topological melanocyte-keratinocyte relationship and melanosome distribution. Confocal laser microscopy images showed that melanocyte dendritic processes are more frequently branched and elongated in senile lentigo skin than in normal skin. Serial transmission electron micrographs showed that dendritic processes extend into intercellular spaces between keratinocytes. Focused ion beam-scanning electron micrographs showed that dendritic processes in senile lentigo encircle adjacent keratinocytes and accumulate large numbers of melanosomes. Moreover, melanosomes transferred to keratinocytes are present not only in the supranuclear area but throughout the perinuclear area except on the basal side. The use of these different microscopic methods helped to elucidate the three-dimensional morphology and topology of melanocytes and keratinocytes in senile lentigo. We show that the localization of melanosomes in dendritic processes to the region encircling recipient keratinocytes contributes to efficient melanosome transfer in senile lentigo.

Torula yeast (Candida utilis)-derived glucosylceramide contributes to dermal elasticity in vitro.

Glucosylceramide (GlcCer) is derived from several plants, such as rice, maize, and wheat, and has been reported to retain moisture by functioning as a barrier between the epidermis and the environment. However, there is insufficient research on the effect of GlcCer on dermal elasticity and wrinkles. In this study, we investigated the effects of torula yeast extract and torula yeast-derived GlcCer on dermal elasticity. We measured cell proliferation, collagen production, and collagen gel contraction using human dermal fibroblasts. Torula yeast extract and torula yeast-derived GlcCer increased dermal fibroblast proliferation and collagen production. Collagen gel contraction was promoted by torula yeast extract and torula yeast-derived GlcCer. These results indicate that GlcCer may affect dermal elasticity. Torula yeast extract and torula yeast-derived GlcCer may contribute to the maintenance of dermal elasticity. PRACTICAL APPLICATIONS: In this study, we found that torula yeast-derived glucosylceramide (GlcCer) has an additional function of improving dermal elasticity. With improved elasticity, skin becomes more resilient, thus preventing wrinkles. GlcCer has already been used in cosmetic products to retain skin moisture. Therefore, torula yeast-derived GlcCer can be expected to have several cosmetic applications.

[A first case report of empyema caused by Tetratrichomonas species in Japan].

Trichomonad is a type of flagellate that parasitizes the human oral cavity, intestine, or vagina. However, respiratory infection is rare. We report a patient with pyothorax related to Tetratrichomonas species, which is the first reported case in Japan. The patient was a 66-year-old female. She consulted the Emergency Outpatient Unit of our hospital with fever, dyspnea, and the retention of pleural effusion. The appearance of the pleural effusion collected by thoracic drainage was brown, fetid pus. On microscopy, Trichomonad was detected in the pleural effusion. On a gene test, Tetratrichomonas sp. was identified. Based on this experience, we propose that non-stained specimens should be examined to detect Trichomonad on a puncture fluid test. (Case report).

[Kinking of the endotracheal tube in a prone patient associated with the inadequate withdrawal maneuver of the Pentax-AWS Airway Scope].

We report a case of intraoperative kinking of an endotracheal tube (ETT) in a prone patient during spine surgery. We postulate that one of the risk factors involved with kinking was the inadequate withdrawal maneuver of Pentax-AWS Airway Scope (AWS). Patient was a 69-year-old woman with hypertension, diabetes mellitus, and rheumatoid arthritis, undergoing C4-6 laminoplasty under general anesthesia in the prone position. A 7.0-mm polyvinyl endotracheal tube (Paker Flex-Tip Tube) was placed to 21 cm at the right angle of the mouse without difficulty using the AWS. Both peak inspiratory pressure (PIP) and partial pressure of end-tidal carbon dioxide began to rise gradually from 24 to 28 cmH2O and 38 to 44 mmHg, respectively. Although over 30 cmH2O in PIP repeatedly appeared after that, we did not find any remarkable change of ventilation except for weak breath sound. Thereafter, when we checked the tube with a flexible fiberoptic bronchoscope, it could not pass through the tube. At first, we asked the surgeon to release neck flexion as much as possible. This procedure could not correct the kink completely but allowed the passage of bronchoscope in the ETT. Then, we tried to reposition the ETT by inserting the bronchoscope beyond the point of kinking for maintaining luminal patency and adequate ventilation. The subsequent anesthetic course was uneventful. Kinking of the ETT in the oral cavity is an uncommon problem but we must keep in mind as one of the differential diagnoses. When using the AWS for endotracheal intubation, we recommend the confirmation of the position of the ETT to be normal in the oral cavity by direct laryngoscopy.

[A case of late drug-eluting stent thrombosis during postoperative period after 5 years of stent implantation].

Stent thrombosis during perioperative period is a critical complication for patients treated with drug-eluting stent (DES). We experienced a case of late DES thrombosis 5 years after initial implantation. A 48-year-old man with familial hyperlipidemia, angina pectoris and chronic pulmonary emphysema, was diagnosed with esophageal carcinoma, and scheduled for esophagectomy. He was first treated with DES 5 years ago, and he underwent repeated revascularization for re-stenosis. He had received anti-platelet therapy up to 1 week prior to the current operation, which was replaced by heparin administration. The surgical procedure was uneventful, and he tolerated it well. Immediately after his admission to ICU, sporadic premature ventricular contraction and ST-elevation occurred, leading to ventricular fibrillation. Emergent coronary angiography revealed re-stenosis of the right coronary artery treated with DES 5 years ago. At present, there was no definite guideline, on the management of DES during perioperative period. It is important for us to decide continuing antiplatelet therapy balancing the risk of stent thrombosis with surgical bleeding in each patient.

[A case of transient postoperative median nerve palsy due to the use of the wrist holder to stabilize an intra-arterial catheter].

We experienced a case of right median nerve palsy at the distal forearm following abdominal surgery. We postulate that the cause of the median nerve palsy is overextension of the wrist by the inappropriate fixation with a holder. The patient was a 46-year-old man with habit of smoking receiving low-anterior resection of the rectum under general and epidural anesthesia in lithotomy position. During surgery his upper limbs were placed on padded arm board abducted about 80 degrees and affixed with soft cotton. His forearms were slightly supinated, whereas his elbows were not over-extended. A 22 G cannula was inserted in the right radial artery and the right wrist was fixed with plastic-holder with soft pad. This position was maintained throughout the operation approximetly for 250 minutes. During anesthesia any special events regarding hemodynamic variables were not observed. He complained of numbness in the palmar side of the digits 1-3 on his right hand without motor disturbance 4 hours after the operation. Examination by the anesthesiologist revealed median nerve palsy. Fortunately, this symptom gradually but completely resolved over the next few days. The possible causes of this neuropathy include the overextension of the wrist or the unexpected extension of the elbow beyond the acceptable range by the supination of forearm, which was induced by the attachment used to stabilize an intra-arterial catheter. Therefore, in the current case we should have returned the wrists promptly to the neutral position following arterial catheter placement to prevent the median nerve palsy. This case suggests the importance of holding the proper position of the arm during surgery.

Topical application of plant extracts containing xanthine derivatives can prevent UV-induced wrinkle formation in hairless mice.

BACKGROUND: Plants are the source of important products with nutritional and therapeutic value. Topical or oral administration of some plant extracts has been shown to reduce photodamage. Cacao bean and cola nut are popular edible plants that contain polyphenols and xanthine derivatives. These plant extracts possess protective effects against UV-induced erythema when taken orally, and an H(2)O(2)-scavenging effect. METHODS: Plant extracts containing xanthine derivatives and three xanthine derivatives were topically applied to the dorsal skin of hairless mice, and the mice were exposed to a resemblance of solar ultraviolet irradiation at a dose of 13.0 J/cm(2) (UVA) for 15 weeks, five times a week on weekdays. After the final irradiation, histological, and analytical studies were performed. RESULTS: Topical application of plant extracts (cacao beans, cola nuts) and caffeine, theobromine, and theophylline markedly prevented photodamage including wrinkle formation and histological alterations. A significant increase in total hydroxyproline content caused by UV irradiation was observed. In contrast, topical application of plant extracts and xanthine derivatives reduced total hydroxyproline and pepsin-resistant hydroxyproline content in comparison with that of the control (vehicle, UV-irradiation group). Moreover, naphthol AS-D chloroacetate esterase staining and diaminobenzidine staining suggested that leukocytes including neutrophils increased in the UV-exposed skin. In contrast, weak staining was observed in skin treated with xanthine derivatives. CONCLUSION: Topical application of plant extracts and xanthine derivatives suppressed wrinkle formation, dermal connective alteration, and collagen accumulation. It is suggested that xanthine derivatives prevented neutrophil infiltration caused by UV-irradiation.

Ergocalciferol promotes in vivo differentiation of keratinocytes and reduces photodamage caused by ultraviolet irradiation in hairless mice.

BACKGROUND: Ergocalciferol (VD(2)) is usually administered orally and it is metabolized to produce its biologically active metabolites in the liver and kidney. Active vitamin D is a well-known potent regulator of cell growth and differentiation. PURPOSE: Active vitamin D such as 1,25-dihydroxyvitamin D(3) (1alpha,25(OH)(2)D(3)) prevents photodamage, including wrinkles and morphologic alterations. However, its clinical and cosmetic use is limited because of its potent, associated effect on calcium metabolism. We examined the efficacy of vitamin D analogues with few adverse effects for preventing skin photodamage. METHOD: Topical application of VD(2) to hairless mouse dorsal skin, and exposure to solar-simulating ultraviolet (UV) radiation at a dose of 10.8 J/cm(2) (UVA) were performed for 15 weeks, five times a week on weekdays. At the end of the final irradiation, histological and analytical studies were performed. RESULTS: Topical application of VD(2) significantly prevented wrinkle formation and abnormal accumulation of extracellular matrix components. In addition, VD(2) suppressed excessive secretion of IL-6 induced by UV irradiation in cultured human normal keratinocytes, in a dose-dependent manner. CONCLUSION: VD(2) promoted keratinocytes differentiation in the epidermis and showed diverse physiological effects, the same as the active form of VD(3). The results suggested that the suppression of skin photodamage involved the promotion of keratinocytes differentiation and suppression of IL-6 secretion induced by exposure to UV. Topical application of VD(2) may become an effective means to suppress solar UV-induced human skin damage.