Paris in the spring: A review of the trade, conservation and opportunities in the shift from wild harvest to cultivation of Paris polyphylla (Trilliaceae).

ETHNOPHARMACOLOGICAL RELEVANCE: P. polyphylla Smith is used in traditional medicine in China, India and Nepal and is likely to be similarly used through most of its geographic range. China is at the centre of demand for P. polyphylla where it is used as an ingredient in several very successful Chinese medicinal herbal formulations. The Chinese e-commerce platform 'alibaba.com', for example, lists 97 P. polyphylla items offered by 46 Asian suppliers, of which 21 are situated in the Chinese mainland, 12 in Nepal, 7 in India, 2 in Pakistan, and 1 each in Bhutan, Hong Kong, Thailand, and Vietnam. Products offered include the crude drug (dried whole or cut rhizomes), extracts and formulations containing this herbal drug. AIMS OF THE REVIEW: The aims of this review were to assess the scale of the P. polyphylla trade, reviewing evidence on the impacts of wild harvest on P. polyphylla populations and on the role of cultivation as an alternative to wild harvest. MATERIALS AND METHODS: Firstly, we reviewed published information on Paris population biology and studies on impacts of wild P. polyphylla harvest from across the geographic range of this species. Secondly, global trade data for P. polyphylla were analysed. Thirdly, we reviewed published information on P. polyphylla cultivation and made field visits to P. polyphylla cultivation areas in Yunnan and Sichuan. RESULTS: Since the 1980s, there has been a 400-fold increase in the market price paid in China for P. polyphylla rhizomes, from 2.7 Chinese Yuan (CNY) per kg in the 1980s to market prices up to 1100 CNY per kg in 2017. Cross-border trade in dried P. polyphylla rhizomes occurs at three different scales. Firstly, an internal, national trade of P. polyphylla rhizomes within countries (such as India, Nepal and China). Secondly, trade in P. polyphylla rhizomes from Nepal (and possibly from Bhutan) to the two range states that have the largest traditional medicine trade in the world: China and India. Thirdly, trade in processed herbal products. In China, for example, P. polyphylla is widely used as an ingredient in several very successful herbal products, including a famous first aid treatment to stop bleeding. Some of these products are exported globally, in addition to entering into regional trade. Trade data in our review shows that c. 800-1050 t of P. polyphylla rhizomes are sold annually, significantly more than recorded in earlier studies. China is the only country where P. polyphylla is cultivated on a significant scale, although small-scale cultivation is taking place in India and Nepal. CONCLUSIONS: Based on the criteria for the inclusion of species in CITES Appendix II (Art. IV 2(a)), there is compelling evidence for adding Paris polyphylla. At the same time, cultivation of P. polyphylla outside of high conservation value habitats needs to be encouraged and supported. One way of doing this may be to develop separate, traceable supply chains for cultivated supplies in order to distinguish them from wild harvested stocks.

Interleukin 4 and prolonged hypoxia induce a higher gene expression of lysyl hydroxylase 2 and an altered cross-link pattern: important pathogenetic steps in early and late stage of systemic scleroderma?

The major pathological processes of systemic scleroderma (SSc) comprise inflammation and microvascular damage in the early or acute progressive stage as well as tissue fibrosis and hypoxia in the chronic end stage. Fibrosis seems to be a general phenomenon characterized by an increase of hydroxylysine aldehyde derived collagen cross-links which has been shown in vitro for systemic scleroderma fibroblasts. In the present study, we analyzed the cross-link pattern and the gene expression of lysyl hydroxylase 2 (LH2) in the skin of SSc. Furthermore, we determined the modulatory impact of inflammatory cytokines (interleukin 4, TNF- alphaand interleukin 1alpha/beta) and prolonged hypoxia on the cross-link profile and the gene expression of LH2, respectively. The concentration of hydroxylysine aldehyde derived cross-links was significantly increased in SSc, while the level of lysine aldehyde derived cross-links was not changed. Accordingly, a marked increase of the transcriptional level of LH2 was found. In long term dermal fibroblast cultures, only interleukin 4 induced an increase of hydroxylysine aldehyde derived cross-links accompanied by a higher gene expression of LH2. Furthermore, prolonged hypoxia induced a marked increase of the mRNA level of LH2 in relation to collagen I. The skin of SSc is characterized by an increase of the transcriptional activity of LH2 leading to an altered cross-link pattern. The changes in the quality of the collagenous matrix can also be obtained in cell culture by the exposure of fibroblasts to interleukin 4 or prolonged hypoxia emphasizing the role of this mediator in the acute and the low oxygen tension in the chronic phase of the disease.

MMP-2, TIMP-2 and MT1-MMP are differentially expressed in lesional skin of melanocytic nevi and their expression is modulated by UVB-light.

BACKGROUND: In malignant melanoma, recent studies have demonstrated an important role of matrix-metalloproteinase 2 (MMP-2), its co-activating enzyme membrane-type matrix-metalloproteinase 1 (MT1-MMP), and the endogenous inhibitor of MMP-2, tissue-inhibitor of matrix metalloproteinase 2 (TIMP-2). Melanocytic nevi are benign neoplasms of the melanocytic lineage, but may exhibit dysplastic features that can be difficult to distinguish from early stage melanoma. As shown in earlier studies, nevi show important morphological and phenotypical changes in response to ultraviolet light (UVB) irradiation. OBJECTIVE: To clarify the role of MMP-2, TIMP-2 and MT1-MMP in UVB-irradiated vs. non-irradiated melanocytic nevi. METHODS: Immunohistochemical comparison of the MMP-2, TIMP-2 and MT1-MMP expression pattern. RESULTS: MMP-2 is expressed by lesional keratinocytes and its expression is up-regulated by UVB-irradiation. MMP-2 expression was not observed in melanocytic cells. TIMP-2, by contrast, is predominantly expressed by melanocytic nevus cells, and its expression is in part down-regulated by UVB-irradiation. MT1-MMP is expressed by basal keratinocytes and to a weaker extent by melanocytic nevus cells. CONCLUSIONS: MMP-2 expression by keratinocytes in nevi probably represents the result of activation of keratinocyte turnover in lesional epidermis. MMP-2 could play a role in the downward movement of junctional nevus cells into the dermis. The reduction of TIMP-2 expression in melanocytic cells by UV-light together with the enhanced expression of MMP-2 in the adjacent epidermis may promote basement membrane degradation. The expression pattern of MT1-MMP in close proximity to epithelial-mesenchymal interfaces underlines the synergistic role of MT1-MMP in this process.

Different pattern of collagen cross-links in two sclerotic skin diseases: lipodermatosclerosis and circumscribed scleroderma.

Changes in the process of cross-linking of collagen molecules are associated with defects in the biomechanical stability of the extracellular matrix. Fibrosis of skin is characterized by an increase in pyridinolines, which are hydroxylysine aldehyde derived cross-links usually absent in healthy skin. In this study, we analyzed cross-links in lipodermatosclerosis and localized scleroderma to address the question whether all the mature cross-links currently characterized are increased in fibrosis in addition to the increase in pyridinolines. As psoralen plus ultraviolet A treatment leads to clinical improvement of fibrotic plaques in localized scleroderma we analyzed the cross-link content in lesional skin after bath psoralen plus ultraviolet A therapy. In skin from patients with localized scleroderma an increase in the total number of mature cross-links was found to be due to an increase in both pyridinolines and dehydro-histidinohydroxymerodesmosine. The concentration of histidinohydroxylysinonorleucine was unchanged. By contrast, the total number of mature cross-links was decreased in lipodermatosclerosis. This decrease was caused by a decrease of lysine aldehyde derived cross-links (dehydro-histidinohydroxymerodesmosine and histidinohydroxylysinonorleucine), whereas the concentration of pyridinolines increased. A decrease in the content of pyridinolines after bath psoralen plus ultraviolet A treatment was found in six out of nine patients with localized scleroderma, which might reflect a remodeling of the extracellular matrix. Our data provide evidence that sclerosis of skin is associated with either an increase in the number of cross-links per molecule of collagen or a change in the molecular nature of the cross-links formed.

[Morphological changes in melanocytic nevi induced by exogenous factors]. / Morphologische Veränderungen in melanozytären Nävi durch exogene Faktoren.

Malignant melanoma is the most important differential diagnosis in both clinical and histologic examination of melanocytic nevi. UV exposure with an erythemogenic dose and mechanical irritation of melanocytic nevi are able to induce reversible morphologic changes which simulate malignant melanoma. These changes are associated with an increased expression of HMB-45 antigen. In addition, an increased proliferation and repair activity is observed after UV exposure. The increased number of melanocytes located in suprabasal layers of the epidermis is accompanied by a change in keratinocyte adhesion molecule expression. The UV light is also able to influence the adhesive properties of melanocytes in vitro. Therefore, both keratinocyte-derived factors and direct influence of UV on the melanocyte system are probably responsible for the morphologic changes. While these exogenously evoked changes are reversible, the potential biologic relevance--especially a possible role in the initiation of the carcinogenesis cascade--requires clarification.

[Severe vitamin D deficiency osteomalacia by coincidence of multiple risk factors]. / Schwere Vitamin-D-Mangel-Osteomalazie durch Koinzidenz mehrerer Risikofaktoren.

HISTORY: An 82-year-old woman was admitted with bilateral clavicular fractures after falling out of bed. She complained of pain in all her bones. She reported having undergone a partial gastrectomy (Billroth II) 40 years ago with subsequent milk intolerance and a dislike of direct sun light. INVESTIGATIONS: Scintigraphy revealed multiple areas of abnormal uptake. Conventional radiography showed in both ulnae transverse zones of increased density with a central translucent band. Laboratory tests demonstrated the constellation of increased bone turnover in secondary hyperparathyroidism with hypocalcaemia (1.49 mmol/l) and hypophyosphataemia (0.62 mmol/l). The 25-hydroxyvitamin D-level was too low to be measured. Ileal crest biopsy indicated osteomalacia. TREATMENT AND COURSE: Serum levels of calcium, alkaline phosphatase and vitamin D became normal after 7 weeks of daily administration of 10,000 IU vitamin D and 1000 mg calcium. The severe pains had subsided after two weeks and no longer required analgesics. CONCLUSION: Multiple areas of increased uptake on skeletal scintigraphy, combined with diffuse bone pain and increased alkaline phosphatase, were at first misinterpreted as extensive skeletal metastases. But the history, at first seemingly of little significance, contained three long-term risk factors for the development of vitamin D and calcium deficiency and thus of osteomalacia, namely partial gastrectomy, milk intolerance and inadequate exposure to sun light. Early recognition of risk factors for bone changes will rapidly lead to the correct diagnosis and immediate treatment of the treatable underlying disease.

Ehlers-Danlos syndrome type VI: lysyl hydroxylase deficiency due to a novel point mutation (W612C).

Ehlers-Danlos syndrome type VI (EDS VI) is a rare autosomal recessively inherited disease of connective tissue. The characteristic symptoms are hyperflexibility of joints and hyperelasticity of skin together with marked scoliosis, ocular manifestations and involvement of the vascular system. The underlying biochemical defect in EDS VI is a deficiency in lysyl hydroxylase (PLOD) activity resulting from mutations in the PLOD gene causing a low hydroxylysine content in various tissues. We found that two out of three patients showed a recently described duplication of about 800 bp in their LH mRNA. In the third patient we identified a new point mutation (2036 G-->C) resulting in a substitution of tryptophan by cysteine in the highly conserved C-terminal region of the enzyme (W612C). In addition, this mutation destroys a restriction site of MwoI. Restriction analysis of the patient's cDNA with MwoI showed the sole occurrence of the mutated transcript, while one allele in his genomic DNA contained the MwoI restriction site. Restriction analysis of the genomic DNA of the unaffected parents displayed a heterozygous loss of the restriction site for MwoI in the mother while the DNA of the father appeared normal. Our study demonstrates that the new point mutation (W612C) in conjunction with a functionless allele, most probably a null allele, for the LH gene may explain the functional deficiencies seen in this patient.

One single erythemagenic UV irradiation is more effective in increasing the proliferative activity of melanocytes in melanocytic naevi compared with fractionally applied high doses.

The effect of a single irradiation with UV light on the expression of Ki67 antigen, topoisomerase II alpha, proliferating cell nuclear antigen (PCNA), the melanocyte activation marker HMB-45 and protein p53 in melanocytic naevi was investigated 1 week after application of a single erythemagenic UV dose and after daily exposures with suberythemagenic doses over 4-6 weeks. To assess the effect of UV irradiation, one half of each naevus was shielded with black tape during the UV exposure, and the irradiated part and the non-irradiated parts were evaluated separately. Except for HMB-45, a double staining procedure was performed to distinguish between labelled melanocytes and keratinocytes. After semiquantitative assessment of the staining signal the irradiated part was compared with the non-irradiated part of the same naevus. Morphological changes and an enhanced proliferative/ reparative activity in melanocytes were much more frequent in the naevi irradiated with a single erythemagenic UV dose than in those given repeated suberythemagenic doses. In addition, the keratinocytes showed an increased labelling for PCNA and p53 after the single irradiation. These data may support the importance of intermittent UV exposure and sunburns in the development of both benign and malignant melanocytic lesions.

Altered x-ray diffraction pattern is accompanied by a change in the mode of cross-link formation in lipodermatosclerosis.

We studied the molecular packing of collagen fibrils by x-ray diffraction in skin specimens of patients with lipodermatosclerosis and in controls. A difference in the tilt angles of the collagen molecules relative to the fiber axis is suggested by a D-stagger that is 1 nm larger in sclerotic skin than in normal skin. In parallel, the collagen cross-links in the skin specimens were analyzed, and a marked increase of both hydroxylysylpyridinoline and lysylpyridinoline, the trivalent mature cross-links characteristic of skeletal tissues, was found. The content of hydroxylysylpyridinoline and lysylpyridinoline was higher in the deep layer of the affected dermis than in the superficial dermis. This increase was always accompanied by an increase in the hydroxylysylpyridinoline/lysylpyridinoline ratio, suggesting that hydroxylysylpyridinoline is a sclerosis-associated cross-link. In addition, lysyl hydroxylation was increased in affected skin, and this increase was apparently restricted to the collagen telopeptides, which are crucial anchoring structures for lysyl dependent cross-links.

Collagen synthesis in (sun-) aged human skin and in fibroblasts derived from sun-exposed and sun-protected body sites.

Endogenous and sun-induced aging of the skin cause distinct morphological alterations. In this study, we have analysed the ratio of collagen III to collagen III plus I in extracts of sun-exposed (face) and sun-protected (abdomen) aged skin, as well as in collagens synthesized by fibroblasts during in vitro culture derived from actinically damaged and sun-protected skin of other subjects (face, medial aspect of the upper arm vs. abdomen, lateral aspect of the forearm). Furthermore, the amount and extent of post-translational modifications of newly synthesized collagens were determined. Chronic sun exposure of the skin does not have an impact on the quantity of collagenous proteins newly synthesized in cell culture. The proportion of collagen III in pepsin extracts of sun-damaged skin is increased relative to sun-protected skin. However, fibroblasts derived from sun-exposed skin synthesize a lower proportion of collagen III than cells from sun-protected skin. The hydroxylation of lysyl residues in newly synthesized alpha 2(I) and alpha 1(III) collagen chains is reduced by UV irradiation, whereas hydroxylation of lysyl residues in alpha 1(I) chains and of prolyl residues in alpha 1(I), alpha 2(I) and alpha 1(III) chains is unaffected by UV irradiation. These data provide circumstantial evidence to indicate that collagen synthesis is influenced independently by endogenous and sun-induced aging.

Attachment of intrinsically and extrinsically aged fibroblasts on collagen and fibronectin.

In the present study we compare human dermal fibroblasts from donors of different age and from sites differing in sun exposure for their capacity to adhere to collagen or fibronectin. Attachment of cells was not dependent on the collagen concentration but was clearly dependent on the fibronectin concentration used for the coating of the plastic surfaces. Attachment of fibroblasts to collagen and fibronectin is dominated by specific integrin binding: only few cells were able to attach to collagen after inhibition with an anti-VLA 2 antibody, or to attach to fibronectin after inhibition with an anti-VLA 5 antibody. On unexposed sites, cells from old donors showed a significantly increased adhesion capacity on collagen (plus 50.7%) and on fibronectin (plus 62.4%) and an increased staining pattern of VLA 2 and VLA 5 integrins in immunohistochemistry in comparison with young donors. In contrast fibroblasts of chronically sun-exposed skin had a significantly decreased adhesion capacity both on collagen (minus 55.3%) and on fibronectin (minus 46.5%) and a poor staining pattern of the above integrins in comparison with cells from solely aged skin (unexposed sites of old donors). Adhesion of all cells could be inhibited by specific integrin antibodies showing that the employed antibodies were able to detect the epitopes responsible for attachment. Intrinsic and extrinsic aging are able to alter cellular properties of mesenchymal cells, such as adhesion to physiologically relevant macromolecules of the extracellular matrix.

Analysis of the age-related composition of human skin collagen and collagens synthesized by fibroblast culture.

Age-related differences in the composition and the post-translational modifications of human skin collagens were examined in the present study. The data were compared with results of collagen synthesis from in vivo-aged fibroblasts in culture. Skin extracts and newly synthesized collagen from fibroblast cultures derived from both old and young donor groups showed the same ratio of collagen III to collagen I. Furthermore, no difference was noted in the degree of prolyl and lysyl hydroxylation of collagen I and collagen III Young and old fibroblasts synthesized a similar quantity of collagen in vitro. The data suggest that fibroblasts maintain a uniform level of collagen production, composition and modification independent of the age of the donor.