Anti-allergic effect of Cyclopia (honeybush) extracts via anti-degranulation activity in a murine allergy model for inhaled antigen.

The anti-allergic effects of extracts prepared from two species of honeybush, Cyclopia genistoides and Cyclopia subternata, were demonstrated in vivo in a murine allergy model for inhaled antigen induced with ovalbumin (OVA) inhalation to mimic pollen allergy. Intake of the extracts increased the production of OVA-specific immunoglobulin (Ig) E (IgE), IgG1, and IgG2a antibodies in serum and significantly suppressed anaphylactic reaction-induced body temperature decline. Moreover, the extracts significantly inhibited antigen-antibody-induced degranulation in RBL-2H3 cells. They also inhibited body temperature decline when the allergic mice were given them after antigen sensitization, indicating that anti-degranulation activity is the major mechanism underlying the anti-allergic effect of Cyclopia extracts. Despite their qualitative and quantitative differences in phenolic composition, the two extracts exhibited similar effects, suggesting that several active compounds might be involved in the activity. Therefore, oral administration of either Cyclopia extract potentially exerts a systemic anti-allergic effect, supporting the increased consumption of honeybush tea for general wellness and improved quality of life.

A male fetus with cyclopia was discovered after miscarriage: A rare case report from Syria.

Key Clinical Message: This case of alobar holoprosencephaly and cyclopia emphasizes the value of prenatal check-ups, particularly in low-income countries. Early ultrasound diagnosis leads to early gestational termination, preventing psychological trauma for the parents. Abstract: Alobar holoprosencephaly is a rare-occurrence malformation with a bad prognosis linked to cyclopia, the most severe cranial feature. Prenatal examinations are essential for identifying these deformities and preventing parental mental health damage.

The effect of Aspalathin linearis, Cyclopia intermedia and Sutherlandia frutescene on sperm functional parameters of healthy male wistar rats.

Introduction: Rooibos (Aspalathin linearis), honeybush (Cyclopia intermedia), and sutherlandia (Sutherlandia frutescene) are three Southern Africa indigenous plants, of which the extracts have become house-hold items and are consumed on a large scale. Although, they are known for their antioxidant properties, studies have highlighted danger in the excessive intake. Therefore, the current study investigated whether treatment with rooibos, honeybush, and sutherlandia will impact sperm functional parameters positively or otherwise, in healthy rats. Methods: Fourteen-week-old pathogen-free adult male Wistar rats (250-300 g) were randomly divided into four groups of ten, including a control, rooibos (RF), honeybush (HB) and a sutherlandia (SL) group. After 7 weeks of treatment, animals were sacrificed. Spermatozoa were retrieved from the cauda epididymis for motility, morphology and concentration analysis and the testis was used for all biochemical assays. Results: The infusion treated animals (RF, HB, and SL) presented with a non-significant decrease of -14.3%, -18.2%, -17.2% and -24.8%, -20.7%, -27.3% in total motility and progressive motility when compared to the control group, respectively. There was a significant increase in number of spermatozoa with slow speed (p = 0.03), especially in SL treated group compared to the control (p = 0.03). Additionally, there was an increase of 28.8%, 31.7%, 23% in superoxide dismutase (SOD) activity of RF, HB and SL compared to control, respectively. This was accompanied with a percentage decrease of -21.1%, -23.7%, 45.9% in malondialdehyde (MDA) levels compared to the control group. Conclusion: In summary, animals treated with the respective infusions presented with a percentage increase in SOD activity but have reduced sperm motility and decreased normal morphology. Paradoxically, they presented with increased sperm concentration. Hence, it is presumed that rooibos, honeybush and sutherlandia may enhance sperm quantity (concentration) but may impair sperm quality (motility morphology) when consumed by healthy animals.

Analysis of the Differentially Expressed Proteins and Metabolic Pathways of Honeybush (Cyclopia subternata) in Response to Water Deficit Stress.

Honeybush (Cyclopia spp.) is a rich source of antioxidant properties and phenolic compounds. Water availability plays a crucial role in plant metabolic processes, and it contributes to overall quality. Thus, this study aimed to investigate changes in molecular functions, cellular components, and biological processes of Cyclopia subternata exposed to different water stress conditions, which include well-watered (as Control, T1), semi-water stressed (T2), and water-deprived (T3) potted plants. Samples were also collected from a well-watered commercial farm first cultivated in 2013 (T13) and then cultivated in 2017 (T17) and 2019 (T19). Differentially expressed proteins extracted from C. subternata leaves were identified using LC-MS/MS spectrometry. A total of 11 differentially expressed proteins (DEPs) were identified using Fisher's exact test (p < 0.00100). Only α-glucan phosphorylase was found to be statistically common between T17 and T19 (p < 0.00100). Notably, α-glucan phosphorylase was upregulated in the older vegetation (T17) and downregulated in T19 by 1.41-fold. This result suggests that α-glucan phosphorylase was needed in T17 to support the metabolic pathway. In T19, five DEPs were upregulated, while the other six were downregulated. Based on gene ontology, the DEPs in the stressed plant were associated with cellular and metabolic processes, response to stimulus, binding, catalytic activity, and cellular anatomical entity. Differentially expressed proteins were clustered based on the Kyoto Encyclopedia of Genes and Genomes (KEGG), and sequences were linked to metabolic pathways via enzyme code and KEGG ortholog. Most proteins were involved in photosynthesis, phenylpropanoid biosynthesis, thiamine, and purine metabolism. This study revealed the presence of trans-cinnamate 4-monooxygenase, an intermediate for the biosynthesis of a large number of substances, such as phenylpropanoids and flavonoids.

Photoprotection and Antiaging Activity of Extracts from Honeybush (Cyclopia sp.)-In Vitro Wound Healing and Inhibition of the Skin Extracellular Matrix Enzymes: Tyrosinase, Collagenase, Elastase and Hyaluronidase.

Cyclopia sp. (honeybush) is an African shrub known as a rich source of polyphenols. The biological effects of fermented honeybush extracts were investigated. The influence of honeybush extracts on extracellular matrix (ECM) enzymes responsible for the skin malfunction and aging process-collagenase, elastase, tyrosinase and hyaluronidase-was analysed. The research also included assessment of the in vitro photoprotection efficiency of honeybush extracts and their contribution to the wound healing process. Antioxidant properties of the prepared extracts were evaluated, and quantification of the main compounds in the extracts was achieved. The research showed that the analysed extracts had a significant ability to inhibit collagenase, tyrosinase and hyaluronidase and a weak influence on elastase activity. Tyrosinase was inhibited effectively by honeybush acetone (IC50 26.18 ± 1.45 µg/mL), ethanol (IC50 45.99 ± 0.76 µg/mL) and water (IC50 67.42 ± 1.75 µg/mL) extracts. Significant hyaluronidase inhibition was observed for ethanol, acetone and water extracts (IC50 were 10.99 ± 1.56, 13.21 ± 0.39 and 14.62 ± 0.21µg/mL, respectively). Collagenase activity was inhibited effectively by honeybush acetone extract (IC50 42.5 ± 1.05 µg/mL). The wound healing properties of the honeybush extracts, estimated in vitro in human keratinocytes (HaCaTs), were indicated for water and ethanol extracts. In vitro sun protection factor (SPF in vitro) showed medium photoprotection potential for all the honeybush extracts. The quantity of polyphenolic compounds was estimated with the use of high-performance liquid chromatography equipped with diode-array detection (HPLC-DAD), indicating the highest mangiferin contents in ethanol, acetone and n-butanol extracts, while in the water extract hesperidin was the dominant compound. The antioxidant properties of the honeybush extracts were estimated with FRAP (2,4,6-Tris(2-pyridyl)-s-triazine) and DPPH (2,2-diphenyl-1-picrylhydrazyl) tests, indicating their strong antioxidant activity, similar to ascorbic acid for the acetone extract in both tests. The wound healing abilities, estimation of SPF in vitro and the direct influence on selected enzymes (elastase, tyrosinase, collagenase and hyaluronidase) of the tested honeybush extracts were analysed for the first time, indicating a high potential of these well-known herbal tea for antiaging, anti-inflammation, regeneration and protection of the skin.

Cyclopia extracts act as selective estrogen receptor subtype downregulators in estrogen receptor positive breast cancer cell lines: Comparison to standard of care breast cancer endocrine therapies and a selective estrogen receptor agonist and antagonist.

Breast cancer is the most diagnosed type of cancer amongst women in economically developing countries and globally. Most breast cancers express estrogen receptor alpha (ERα) and are categorized as positive (ER+) breast cancer. Endocrine therapies such as, selective estrogen receptor modulators (SERMs), aromatase inhibitors (AIs), and selective estrogen receptor downregulators (SERDs) are used to treat ER+ breast cancer. However, despite their effectiveness, severe side-effects and resistance are associated with these endocrine therapies. Thus, it would be highly beneficial to develop breast cancer drugs that are as effective as current therapies, but less toxic with fewer side effects, and less likely to induce resistance. Extracts of Cyclopia species, an indigenous South African fynbos plant, have been shown to possess phenolic compounds that exhibit phytoestrogenic and chemopreventive activities against breast cancer development and progression. In the current study, three well characterized Cyclopia extracts, SM6Met, cup of tea (CoT) and P104, were examined for their abilities to modulate the levels of the estrogen receptor subtypes, estrogen receptor alpha and estrogen receptor beta (ERß), which have been recognized as crucial to breast cancer prognosis and treatment. We showed that the Cyclopia subternata Vogel (C. subternata Vogel) extracts, SM6Met and cup of tea, but not the C. genistoides extract, P104, reduced estrogen receptor alpha protein levels while elevating estrogen receptor beta protein levels, thereby reducing the ERα:ERß ratio in a similar manner as standard of care breast cancer endocrine therapies such as fulvestrant (selective estrogen receptor downregulator) and 4-hydroxytamoxifen (elective estrogen receptor modulator). Estrogen receptor alpha expression enhances the proliferation of breast cancer cells while estrogen receptor beta inhibits the proliferative activities of estrogen receptor alpha. We also showed that in terms of the molecular mechanisms involved all the Cyclopia extracts regulated estrogen receptor alpha and estrogen receptor beta protein levels through both transcriptional and translational, and proteasomal degradation mechanisms. Therefore, from our findings, we proffer that the C. subternata Vogel extracts, SM6Met and cup of tea, but not the C. genistoides extract, P104, selectively modulate estrogen receptor subtypes levels in a manner that generally supports inhibition of breast cancer proliferation, thereby demonstrating attributes that could be explored as potential therapeutic agents for breast cancer.

Fetal Cyclopia, Proboscis, Holoprosencephaly, and Polydactyly: A Case Report With Review of Literature.

We present a case of holoprosencephaly (HPE) with cyclopia and proboscis. The mother was a 35-year-old, G1P1 with no known comorbid conditions, not in a consanguineous marriage, and with no history of illicit drug use. On a routine antenatal ultrasound scan, features of alobar HPE, proboscis, and other anomalies were identified. The mother was counseled about the condition and as per their consent, the pregnancy was terminated. After labor induction, she gave birth to a female neonate weighing 1,000 g. The newborn's Apgar score could not be calculated. In the initial physical examination, an eye and a 3.5-cm proboscis were seen in the middle of the forehead. The newborn had no nose, and the outer ears were normal. On postmortem examination, alobar HPE, polydactyly, ventricular septal defect, and myelomeningocele were confirmed. This case report highlights the importance of attention to these details during antenatal scans for early detection in order to reduce the maternal and neonatal health burden. The pictures presented in this article were taken after obtaining parental consent.

Rare manifestations of alobar holoprosencephaly and the potential causes: a report of two cases.

Holoprosencephaly is a rare and possibly fatal neural tube defect represented by complete or partial forebrain noncleavage. It can be classified into four types: alobar, semilobar, lobar, and middle interhemispheric fusion variant. It is usually diagnosed through prenatal ultrasound or after birth by visually observing the morphological abnormalities and/or through neurological screening. Potential causes include maternal diabetes, alcoholism, infections during pregnancy, drugs, and genetic causes. Case presentation: Herein, we report two cases of holoprosencephaly's rarest manifestations, albeit cebocephaly in the first case, and cyclopia with a probocis in the second. Cebocephaly, (hypotelorism with a single nostril and a blind-ended nose) was present in the first case; a Syrian newborn girl for a 41-year-old mother who works in collecting Capparis spinosa, and cyclopia with skull vault absence and posterior encephalocele in the second case; a Syrian newborn girl for a 26-year-old mother, the parents here where second-degree relatives. Conclusions: Early diagnosis through ultrasound is preferred in such cases and management options should be assessed and discussed with the parents due to poor prognosis. Adherence to pregnancy follow-up programs is essential to detect malformations and disorders as early as possible, especially when risk factors exist. Also, this paper may suggest a potential correlation between C. spinosa and holoprosencephaly. Therefore, we suggest that more research should be done.

Pericentric Inversion of Chromosome 9 in Twins With Cyclopia: A Rare Entity.

Pericentric inversion of chromosome 9 (inv(9)) is one of the most common variants seen in a normal human karyotype that occurs during meiosis. Despite being categorized as a normal variant, some studies using classical cytogenetics have recently shown that inv(9) could be associated with azoospermia, congenital anomalies, growth retardation, and rarely with abnormal karyotype. However, there is no reported association with cyclopia. Interestingly this genetic variant involves twin fetuses. A 36-year-old multiparous lady with dichorionic diamniotic twin pregnancy presented to the fetomaternal unit with fetal growth restriction at 34 weeks of gestation. An ultrasound scan revealed both have microcephaly, fisting hands, holoprosencephaly, and proboscis suspicious of Patau syndrome. Amniocentesis was not issued due to late pregnancy and guarded prognosis. The mother presented with pre-eclampsia at 35 weeks of gestation. The pregnancy managed to prolong up to 36 weeks after which caesarean section was performed due to the leading twin being in a transverse lie. Two baby twin girls were born 3 minutes apart with microcephaly and cyclops appearance. Chromosomal analysis of both twins revealed similar karyotypes of 46, XX, inv(9)(p11,q13). Pericentric inversion of chromosome 9 is regarded as a normal chromosomal variation in the general population, but in twins with cyclops is considered rare. Early referral to a tertiary hospital for twin management is highly required. It may identify fetuses with such abnormalities and counsel the parents with appropriate management.

Cyclopia intermedia (Honeybush) Induces Uncoupling Protein 1 and Peroxisome Proliferator-Activated Receptor Alpha Expression in Obese Diabetic Female db/db Mice.

Previously, we reported that a crude polyphenol-enriched fraction of Cyclopia intermedia (CPEF), a plant consumed as the herbal tea, commonly known as honeybush, reduced lipid content in 3T3-L1 adipocytes and inhibited body weight gain in obese, diabetic female leptin receptor-deficient (db/db) mice. In the current study, the mechanisms underlying decreased body weight gain in db/db mice were further elucidated using western blot analysis and in silico approaches. CPEF induced uncoupling protein 1 (UCP1, 3.4-fold, p < 0.05) and peroxisome proliferator-activated receptor alpha (PPARα, 2.6-fold, p < 0.05) expression in brown adipose tissue. In the liver, CPEF induced PPARα expression (2.2-fold, p < 0.05), which was accompanied by a 31.9% decrease in fat droplets in Hematoxylin and Eosin (H&E)-stained liver sections (p < 0.001). Molecular docking analysis revealed that the CPEF compounds, hesperidin and neoponcirin, had the highest binding affinities for UCP1 and PPARα, respectively. This was validated with stabilising intermolecular interactions within the active sites of UCP1 and PPARα when complexed with these compounds. This study suggests that CPEF may exert its anti-obesity effects by promoting thermogenesis and fatty acid oxidation via inducing UCP1 and PPARα expression, and that hesperidin and neoponcirin may be responsible for these effects. Findings from this study could pave the way for designing target-specific anti-obesity therapeutics from C. intermedia.

Co-Treatment of Caco-2 Cells with Doxorubicin and Gold Nanoparticles Produced from Cyclopia intermedia Extracts or Mangiferin Enhances Drug Effects.

Mangiferin (MGF) is a natural and valuable polyphenol found in significant levels in many plant species, including Cyclopia intermedia (C. intermedia). In a previous study, we synthesized gold nanoparticles (AuNPs) using MGF and a water extract of C. intermedia and reported that these AuNPs have very low cytotoxicity toward a human colon cancer (Caco-2) cell line. Although the study also showed that these biogenic AuNPs in combination with doxorubic (DOX) significantly augmented the cytotoxic effects of DOX in Caco-2 cells, the mechanism of the enhanced effect was not fully understood, and it was also not known if other cell lines would be sensitive to this co-treatment. In the present study, we examined the cytotoxicity of the co-treatment in Caski, HeLa, HT-29, KMST-6 and MDA-321 cell lines. Additionally, we investigated the mechanistic effects of this co-treatment in Caco-2 cells using several assays, including the adenosine triphosphate (ATP), the oxidative stress, the mitochondrial depolarization, the colony formation, the APOPercentage and the DNA fragmentation assays. We also assessed the intracellular uptake of the biogenic AuNPs. The study showed that the biogenic AuNPs were effectively taken up by the cancer cells, which, in turn, may have enhanced the sensitivity of Caco-2 cells to DOX. Moreover, the combination of the biogenic AuNPs and DOX caused a rapid depletion of ATP levels, increased mitochondrial depolarization, induced apoptosis, reduced the production of reactive oxygen species (ROS) and inhibited the long-term survival of Caco-2 cells. Although the study provided some insight into the mechanism of cytotoxicity induced by the co-treatment, further mechanistic and molecular studies are required to fully elucidate the enhanced anticancer effect of the co-treatment.

Combinatorial treatments of tamoxifen and SM6Met, an extract from Cyclopia subternata Vogel, are superior to either treatment alone in MCF-7 cells.

Synergistic drug combinations are not only popular in antibiotic, anti-microbial, immune disease (i.e., AIDS) and viral infection studies, but has also gained traction in the field of cancer research as a multi-targeted approach. It has the potential to lower the doses needed of standard of care (SOC) therapeutic agents, whilst maintaining an effective therapeutic level. Lower dosages could ameliorate the fundamental problems such as drug resistance and metastasis associated with current SOC therapies. In the current study, we show that the combination of SM6Met with (2)-4-hydroxytamoxifen (4-OH-Tam, the active metabolite of tamoxifen) produces a strong synergistic effect in terms of inhibiting MCF7 ER-positive (ER+) breast cancer cell proliferation and that a 20 times lower dose of 4-OH-Tam in combination with SM6Met is required to produce the same inhibitory effect on cell proliferation as 4-OH-Tam on its own. Cell cycle analyses of the best combination ratios of SM6Met and 4-OH-Tam also suggests that the combination results in increased accumulation of cells in the S-phase and in the apoptotic phase. Moreover, the best combination ratio (20:1) of SM6Met with 4-OH-Tam displayed greater anti-metastatic potential in terms of inhibiting ER+ breast cancer cell migration, invasion, and colony formation than the SOC therapy alone, suggesting that SM6Met together with 4-OH-Tam could be a viable drug combination for not only delaying resistance and ameliorating the negative side-effects associated with current SOC therapies, like tamoxifen, but could also provide a novel, more affordable therapeutic alternative for treating or preventing ER+ breast cancer metastasis.

Synophtalmia on a newborn.

Synophtalmia or cyclopia is a rare presentation of alobar holoprosencephaly. Cases which have been reported are stillborn or dead in post-delivery period. We are presenting a 3000-g live full-term newborn girl delivered by caesarean section with a well-marked cyclopia, but who died 30 min post-delivery. The case did not present with other abnormalities. The literature showed that genetic disorders are associated with cyclopia. A prenatal anomaly scan can help in the early detection of the condition and timely termination of the pregnancy can be conducted.

First report of Neofusicoccum australe causing dieback of honeybush in the Western Cape, South Africa.

Honeybush (Cyclopia spp.) is an indigenous, leguminous member of the Cape fynbos biome growing in the coastal winter rainfall districts of the Western and Eastern Cape Provinces of South Africa (Joubert et al. 2011). Honeybush is used for the production of herbal teas and is harvested from wild-growing and cultivated plantations (du Toit et al. 1998). Very little is known regarding diseases caused by pathogens on this indigenous plant. Only one report of twig dieback on honeybush caused by several Diaporthe Nitschke species have been reported in South Africa (Smit et al. 2021). Several honeybush producers reported poor growth and dieback in their C. subternata plantations in the Western Cape Province, South Africa. Symptoms included twig dieback, branch dieback, death of branches as well as death of entire plants. In April 2008, branches from 8-year-old cultivated plants with dieback symptoms were collected in Stellenbosch. Fungal isolations were carried out from affected material as described by Van Niekerk et al. (2004) which consistently revealed the presence of a Botryosphaeriaceae species. Two isolates were grown on water agar with sterile pine needles and incubated at 25˚C using a 12-hour day/night cycle and near-ultraviolet light. Pycnidia formed after two weeks. Morphological characteristics similar to Neofusicoccum australe (Slippers, Crous & Wingfield) Crous, Slippers & Phillips were observed (Phillips et al. 2013). Conidia were hyaline, aseptate, fusiform with subtruncate bases (16.8-)18.8-22.1(-24.6) × (4.8-)5.3-6.1(-6.4) µm (n=50). Conidiogenous cells were holoblastic, hyaline and subcylindrical to flask-shaped tapering to the apex (11-15 × 2 µm) (n=10). Colonies on potato dextrose agar were light primrose turning olivaceous grey after 7 days with a light-yellow pigment diffusing into the medium. Mycelia was moderately dense with an appressed centre mat. The identity of the isolates was further confirmed by sequencing the ribosomal RNA Internal Transcribed Spacer (ITS) and the elongation factor 1-alpha (EF-1α) gene regions using primer pairs ITS4-ITS5 (White et al. 1990) and EF1-728F-EF1-986R (Alves et al. 2008), respectively. Sequences had a 100% similarity to N. australe ex-type CMW6837 isolate (accessions AY339262 and AY339270) (Slippers et al. 2004). Two isolates (STEU6554 and STEU6557) were deposited in the culture collection at the Department of Plant Pathology at Stellenbosch University and the sequences were submitted to GenBank with accession numbers ON745603, ON745604, ON746573 and ON746574. Pathogenicity tests using the two N. australe isolates were conducted by inoculating two shoots each of three field-grown C. subternata plants with a 4mm colonised potato dextrose agar (PDA) mycelium plug of each isolate on wounds made by a 4mm cork borer (Van Niekerk et al. 2004). A third shoot was inoculated with a uncolonized PDA plug as the negative control. After 12 weeks, brown-black lesions that were significantly longer (average 55.2 mm) than the uncolonized agar plug control (16.1 mm) were observed. Lesions were observed in all three plants. Neofusicoccum australe was re-isolated (van Niekerk et al. 2004) from all inoculated shoots confirming Koch's postulates. The economic impact and damages caused by N. australe as well as its incidence and severity on honeybush in South Africa is unknown. However, the pathogen caused dieback of entire branches and death of plants indicating that it could be an important pathogen of honeybush. Additionally, N. australe is one of the most important disease-causing Botryosphaeriaceae pathogens on a wide range of economical fruit and vine crops globally (Mojeremane et al. 2020). This is the first report of N. australe as a known pathogen causing decline and dieback of C. subternata in South Africa. References: Alves, A. et al. 2008. Fungal Divers. 28:1. du Toit, J. et al. 1998. J. Sustain. Agric. 12:67. Joubert, E. et al. 2011. S. Afr. J. Bot. 77:887. Mojeremane, K. et al. 2020. Phytopathol. Mediterr. 59:581. Phillips, A. J. et al. 2013. Stud. Mycol. 76:51. Slippers, B. et al. 2004. Mycologia 96:1030. Smit, L. et al. 2021. Eur. J. Plant Pathol. 161:565. van Niekerk, J. M. et al. 2004. Mycologia 96:781. White, T. J. et al. 1990. Pages 315 in: In PCR Protocols: A Guide to Methods and Applications. Academic Press Inc, USA. Declaration. The author(s) declare no conflict of interest Acknowledgments. This work benefitted from the financial support of the Agricultural Research Council, Infruitec-Nietvoorbij, South Africa.

Heat treatment improves the sensory properties of the ultrafiltration by-product of honeybush (Cyclopia genistoides) extract.

BACKGROUND: Ultrafiltration of green honeybush (Cyclopia genistoides) extract results in a by-product (retentate). Application of further separation processes for recovery of polyphenols would entail creation of additional waste. Repurposing the retentate as a food flavour ingredient provides an alternative valorization approach. RESULTS: The retentate, suspended in water (270 g L-1 ), was heat-treated at 80 °C for 2, 4, 8 and 16 h, and at 90 °C for 2, 4, 6 and 8 h to change its sensory profile. The heat-treated retentate, diluted to beverage strength (2.15 g L-1 ), had prominent 'grape/Muscat-like' and 'marmalade/citrus' aroma and flavour notes. Overall, heating for ≤ 4 h increased the intensities of positive flavour and aroma notes, while reducing those of 'green/grass', 'hay' and bitterness, whereafter further heating only had a slight effect on the aroma profile at 80 °C (P < 0.05), but not at 90 °C (P ≥ 0.05). The heat treatments, 80 °C/4 h and 90 °C/4 h, were subsequently applied to different batches of retentate (n = 10) to accommodate the effect of natural product variation. Heating at 90 °C produced higher intensities of positive aroma attributes (P < 0.05), but was more detrimental to the phenolic stability, compared to 80 °C. CONCLUSION: After heat treatment, the phenolic content of C. genistoides retentate, reconstituted to beverage strength, still fell within the range of a typical 'fermented' (oxidized) honeybush leaf tea infusion. The change in phenolic composition will not diminish the benefit of an improved sensory profile for the retentate by-product through heating. © 2021 Society of Chemical Industry.

Cyclopia, a newborn with a single eye, a rare but lethal congenital anomaly: A case report.

INTRODUCTION AND IMPORTANCE: Cyclopia is a rare congenital disorder characterized by facial abnormalities. In this condition, the orbits of the eye are not properly divided into two cavities so that they can be seen either as a single eye field or two bilateral fields that are very close to each other. This syndrome affects the embryos that are either aborted or stillborn upon delivery or, at best, die shortly after birth. CASE PRESENTATION: This case report is of a 37-week- and 5-day-old female fetus with a birth weight of 2300 g, a height of 43 cm, and a head circumference of 31 cm. She was born to a 44-year-old mother through normal vaginal delivery at Mahzad Hospital, Urmia, Iran. In the physical examination, an eye and a 4-cm proboscis were seen in the middle of the forehead. The newborn also had no nose, and his outer ears were normal. No cleft lip or cleft palate was observed. Unfortunately, the newborn expired 13 h after birth. CLINICAL DISCUSSION: Cyclopia leads to a stillbirth since the brain and other parts of the body do not grow normally in fetuses with this disorder. Moreover, it can be diagnosed using ultrasonography while the fetus is growing inside the uterus. CONCLUSION: Early diagnosis during pregnancy using diagnostic methods and proper management of this abnormality should be emphasized to prevent further harm to the newborn and the mother with this syndrome. Moreover, many of these newborns should be offered early neonatal palliative care.

Cyclopia: The Face Predicts the Future.

The most extreme form of holoprosencephaly (HPE) is cyclopia and appears with a single characteristic midline diamond-shaped orbital structure and various facial, brain, and extrafacial features. We aimed to report a case of a cyclopic fetus diagnosed at the 22 weeks of the gestational age and further we reviewed the recent literature in order to highlight the etiopathogenesis and set goals for approaching such future pregnancies. Following the first-trimester assessment, in a 27-year-old pregnant woman, who underwent in vitro fertilization, the pregnancy was associated with a low risk for aneuploidies and a high risk for pre-eclampsia. On the anomaly scan, due to severe fetal brain maldevelopment and microcephale, HPE was suspected. Furthermore, three-dimensional ultrasound confirmed a common orbit in the midline of the face. Although the parents did not opt for amniocentesis and further postnatal management, parental karyotyping test did not detect any pathology. The pregnancy was terminated and the macroscopic examination of the aborted specimen revealed cyclopia, synophalmia, fussed eyelids with a proboscis on the upper midline of the face, and a malpositioned left ear. To conclude, cyclopia is not widely manifested, and different cyclo-pian disorders could still occur. Although this rare congenital abnormality is incompatible with life, the awareness of the spectrum of sonographic features and the appropriate genetic counseling can determine the outcome of current and forthcoming pregnancies.

Alobar holoprosencephaly in mining-related areas of the Eastern region of the Democratic Republic of the Congo: A case series.

BACKGROUND: Recent case reports described three cases of holoprosencephaly (HPE) in the area with high mining-related pollution of the southern region of the Democratic Republic of the Congo (DRC). We reported two male neonates with clinically diagnosed HPE in the localities of Fizi and Kitutu, two mineral areas in the Eastern region of the same country (DRC), where artisanal surface mining is predominant with high exposure to radiation and heavy metals from mining. CASES' PRESENTATIONS: Two newborns from adult and multigravida mothers without pregnancy complication. The birth weights were 3,200 g and 2,500 g, respectively, and the malformations noticed were essentially the single median eye, the absent nose, polydactyly for one case, and proboscis for the other case. They both died a few minutes later after birth. CONCLUSION: The etiologic factors of HPE remain unknown but seem to be multifactorial from both genetic and environmental factors. We hypothesize under reserve for these two cases that mining and radiation expositions were likely potential environmental associated factors to the occurrence of these malformations.

Exposure to ethanol leads to midfacial hypoplasia in a zebrafish model of FASD via indirect interactions with the Shh pathway.

BACKGROUND: Gene-environment interactions are likely to underlie most human birth defects. The most common known environmental contributor to birth defects is prenatal alcohol exposure. Fetal alcohol spectrum disorders (FASD) describe the full range of defects that result from prenatal alcohol exposure. Gene-ethanol interactions underlie susceptibility to FASD, but we lack a mechanistic understanding of these interactions. Here, we leverage the genetic tractability of zebrafish to address this problem. RESULTS: We first show that vangl2, a member of the Wnt/planar cell polarity (Wnt/PCP) pathway that mediates convergent extension movements, strongly interacts with ethanol during late blastula and early gastrula stages. Embryos mutant or heterozygous for vangl2 are sensitized to ethanol-induced midfacial hypoplasia. We performed single-embryo RNA-seq during early embryonic stages to assess individual variation in the transcriptional response to ethanol and determine the mechanism of the vangl2-ethanol interaction. To identify the pathway(s) that are disrupted by ethanol, we used these global changes in gene expression to identify small molecules that mimic the effects of ethanol via the Library of Integrated Network-based Cellular Signatures (LINCS L1000) dataset. Surprisingly, this dataset predicted that the Sonic Hedgehog (Shh) pathway inhibitor, cyclopamine, would mimic the effects of ethanol, despite ethanol not altering the expression levels of direct targets of Shh signaling. Indeed, we found that ethanol and cyclopamine strongly, but indirectly, interact to disrupt midfacial development. Ethanol also interacts with another Wnt/PCP pathway member, gpc4, and a chemical inhibitor of the Wnt/PCP pathway, blebbistatin, phenocopies the effect of ethanol. By characterizing membrane protrusions, we demonstrate that ethanol synergistically interacts with the loss of vangl2 to disrupt cell polarity required for convergent extension movements. CONCLUSIONS: Our results show that the midfacial defects in ethanol-exposed vangl2 mutants are likely due to an indirect interaction between ethanol and the Shh pathway. Vangl2 functions as part of a signaling pathway that regulates coordinated cell movements during midfacial development. Ethanol exposure alters the position of a critical source of Shh signaling that separates the developing eye field into bilateral eyes, allowing the expansion of the midface. Collectively, our results shed light on the mechanism by which the most common teratogen can disrupt development.